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2021 December
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医歯薬出版 ● 新刊・好評書のご案内

世界基準の臨床歯内療法 第 2 版
石井

宏

編著

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この 1 冊で臨床歯内療法のベーシックからアドバンスまでを網羅．
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牛窪敏博・神戸良編著

バイオセラミックマテリアルをより臨床応用しやすいよう，使用方法のみならず，どのようなときになぜ有用なのか，従来法との違い，使用時の注意点などをエビデンスをもとに解説．
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■ A4判変型／72頁／カラー

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■ A4判変型／172頁／カラー■ 定価 8,800円（本体 8,000円＋税10％）

SBN978-4-263-44533-4

OPERATIVE DENTISTRY, ENDODONTOLOGY AND PERIODONTOLOGY
Vol. 1, No. 1
Preface
Launch of the ODEPCONTENTSDECEMBER 2021

TANI-ISHII Nobuyuki

Original Articles
Influence of Etching Mode on the Dentin Bond Effectiveness of a Universal Adhesive
Jennifer A. HASSLEN, Akimasa TSUJIMOTO, Mark D. MARKHAM,
Mone SHOJI, Toshiki TAKAMIZAWA, Mark A. LATTA, Masashi MIYAZAKI,Atsushi KAMIMOTO and Wayne W. BARKMEIER （ 1 ）Development of Chairside Dry Heat Sterilizer Using Halogen Lamps
as a Thermal Radiation Source
Keisuke SAIGUSA, Yukihiro TAKAHASHI, Munehiro MAEDA
and Masaru IGARASHI （ 9 ）Development of Multilayered Dental Pulp Cell Sheets
and Consideration for Calcification Ability
Katofumi KOYANAGI, Munehiro MAEDA, Miki SEKIYA,
Taro NISHIDA and Masaru IGARASHI （ 22 ）Inhibitory Mechanisms of S-PRG Eluate and S-PRG Filler
against Volatilization of Hydrogen Sulfide
Sami OMAGARI, Nao TANIGUCHI, Yoshiyuki YOKOGAWA,
Masahiro YONEDA, Shigeru YAMAMOTO, Takashi HANIOKA
and Takao HIROFUJI （ 30 ）Clinical Assessment of Resin-coating Technique for Dentin after Cavity Preparation
Hanemi TSURUTA, Shusuke KUSAKABE, Michael F BURROW
and Toru NIKAIDO （ 37 ）The Shaping Advantage of M-wire Compared with Conventional Nickel-titanium Rotary Instruments
in Heavy Curvature Canals
Akito KASAHARA, Mika TANAKA-SATO, Noriko MUTOH and Nobuyuki TANI-ISHII （ 44 ）Influence of Surface Moisture on the Bond Strength of a Universal Adhesive in the Etch-and-rinse Mode
Runa SUGIMURA, Akimasa TSUJIMOTO, Kei IWASE,
Shun KATSUKI, Ahmad ALKHAZALEH, Olajide OBE,
Toshiki TAKAMIZAWA and Masashi MIYAZAKI （ 52 ）Removal of Biofilm from Titanium Surfaces Using 400-kHz UltrasoundTakashi TAKIGUCHI, Junki YAMADA, Masanori SATO
and Matsuo YAMAMOTO （ 59 ）Effect of Root Canal Transportation by Minimally Invasive Endodontic Shaping of Canal Orifice Dentin
Kazuyuki TSUBAKI, Mai UTSUNOMIYA, Kaori SHIMOJIMA,
Noriko MUTOH and Nobuyuki TANI-ISHII （ 69 ）Study on Evaluation of Cavity Preparation Using Non-contact High-speed 3-D Shape Measuring Device
Saeko OKUMURA, Hiroaki TANIMOTO, Kenzo YASUO, Naohiro IWATA,
Kazushi YOSHIKAWA and Kazuyo YAMAMOTO （ 77 ）Effects of Universal Adhesives on Dentin Adhesive Performance and Acid-base Resistance
Ryuta ANDO, Takaaki SATO, Naoko MATSUI, Masaomi IKEDA,
Noriko HIRAISHI, Tomohiro TAKAGAKI, Toru NIKAIDO,
Junji TAGAMI and Yasushi SHIMADA （ 88 ）Case Report
A Case of Non-surgical Root Canal Treatment Using an Operating Microscope on Type Ⅲ-b Dens Invaginatus
Masashi YAMADA, Kento ODAKA, Norio KASAHARA,
Satoru MATSUNAGA, Yoshiki TAMIYA, Ryo SAKO
and Masahiro FURUSAWA （ 97 ）Submission Guidelines and Guidance

Publishedby
THE JAPANESE SOCIETY OF CONSERVATIVE DENTISTRY（JSCD）
c/o Oral Health Association of Japan（Kōkūhoken kyōkai）
1‒43‒9, Komagome, Toshima-ku, Tokyo 170‒0003
Japan（ 105 ）

Editorial Board of Operative Dentistr y, Endodontology and Periodontology
Editor-in-Chief
Hideki SHIBA

Graduate School of Biomedical and Health Sciences, Hiroshima University

Associate Editor
Hidefumi MAEDA

Kyushu University Faculty of Dental Science

Editorial Board

Atsushi KAMEYAMA
Matsumoto Dental University

Kazuo KITAMURAThe Nippon Dental University

Satoshi HIRAYAMA
Nihon University School of Dentistry at Matsudo

Takashi MURAMATSU
Tokyo Dental College

Takashi YAEGASHI
School of Dentistry, Iwate Medical University

Kazuyo YAMAMOTO
Osaka Dental University

Matsuo YAMAMOTO
Showa University School of Dentistry

Hiromichi YUMOTO
Tokushima University Graduate School of Biomedical Sciences

Kunihiko YOSHIBA
Niigata University Graduate School of Medical and Dental Sciences

Atsutoshi YOSHIMURA
Nagasaki University Graduate School of Biomedical Sciences

Masahiro YONEDA
Fukuoka Dental College

Editorial Secretar y
Katsuhiro TAKEDA

Graduate School of Biomedical and Health Sciences, Hiroshima University

PREFACE
Launch of the ODEPTANI-ISHII Nobuyuki
President
The Japanese Society of Conservative Dentistry

The Japanese Society of Conservative Dentistry is pleased to announce that it will start publishing its English journal, Operative Dentistry, Endodontology and Periodontology（ODEP）,
from December 2021.
Since the first edition of our journal was published in Japanese in 1958, it has been issued six
times a year since 1986, and 280 volumes have been published to date. The circulation of eachcopy is about 4,500, and these have been mailed to academic society members, but since 2020, it
has been published online and can be distributed to researchers overseas via the Web.
By publishing an English journal, the members of the Japanese Society of Conservative Dentistry will be able to disseminate their scientific research, clinical research, and education in thefield of conservative dentistry to researchers all over the world. It will be a great honor andpleasure for us to contribute to the development of research.
Publication of this ODEP journal will help establish close cooperation with related academic
societies in Japan and overseas；we will strive to conduct research, clinical practice, educationand training that will be disseminated to the world as well as contribute to Japanese dentistry.
I hope that the journal will provide a platform for the further development of conservative
dentistry based on the long-held philosophy of our Society.

Dec, 2021

1

Original Article

ODEP 1（1）：1～8，2021

Influence of Etching Mode on the Dentin Bond Effectivenessof a Universal Adhesive
Jennifer A. HASSLEN1, Akimasa TSUJIMOTO2, Mark D. MARKHAM1,
Mone SHOJI3, Toshiki TAKAMIZAWA3, Mark A. LATTA1,
Masashi MIYAZAKI3, Atsushi KAMIMOTO4 and Wayne W. BARKMEIER1
1

Department of General Dentistry, Creighton University School of Dentistry2

Department of Operative Dentistry, Iowa University School of Dentistry3

Department of Operative Dentistry, Nihon University School of Dentistry4

Department of Comprehensive Dentistry and Clinical Education, Nihon University School of Dentistry

Abstract
Purpose: This laboratory study aimed to assess the dentin bond fatigue resistance and interfacial characteristics of a universal adhesive in etch-and-rinse and self-etch modes.
Methods: The universal adhesive used was Clearfil Universal Bond Quick（Kuraray Noritake Dental,
Tokyo, Japan）. The dentin surfaces were treated with the universal adhesive in either etch-and-rinse or selfetch mode. Initial bond strength was measured by loading 15 specimens per test group to failure using an
all-electric dynamic instrument with a chisel-shaped metal load at a crosshead speed of 1.0 mm/min. The
fatigue load was applied to the specimens using a sine wave at a frequency of 20 Hz for 50,000 cycles or until
failure occurred. The surface free energy characteristics of adhesive-treated dentin were determined. Scanning electron microscopy（SEM）observations of the resin-dentin interface of the adhesive were also conducted for each etching mode.
Results: For the universal adhesive tested, etching mode did not affect the bond fatigue resistance. The
surface free energy characteristics of the baseline showed that the γs, γsd, γsp, and γsh values for dentin in
etch-and-rinse mode were significantly lower than in self-etch mode. The surface free energy values of adhesive-treated dentin in etch-and-rinse mode were not significantly decreased by adhesive application, whileboth γs and γsh values were decreased after adhesive application in self-etch mode. In SEM observations ofadhesive interfaces, deeper penetration of adhesives into the dentinal tubules was observed in etch-and-rinsemode due to removal of the smear layer and opening of the dentinal tubules.
Conclusion: The results suggest that the dentin bonding performance of the universal adhesive was notaffected by etching mode.
Key words: universal adhesive, dentin bonding performance, etching mode

Corresponding author: Akimasa TSUJIMOTO, Department of Operative Dentistry, Iowa University School of Dentistry, S248
Dental Science Building, 801 Newton Rd, Iowa City, IA52422, USA
TEL: ＋1319‒462‒4204, E-mail: akimasa-tsujimoto@uiowa.edu
Received for Publication: November 15, 2020/Accepted for Publication: January 12, 2021
DOI: 10.11471/odep.2021‒001

ODEP Vol. 1, No. 1

2

treated with universal adhesive in the two different
modes may provide an explanation for the discrepancy

Introduction

between bond strength and chemical bonding effectiveRecently, universal adhesives have become popular in
dentistry because they can be used in a variety ofness, as long as such an assessment is accompanied bya bond fatigue resistance analysis.

modes: etch-and-rinse, self-etch, or selective-etch1）. With

The purpose of this laboratory study was to assess

the expiration of the patent for 10-methacryloy-

the dentin bond fatigue resistance and energy charac-

loxydecyl dihydrogen phosphate（10-MDP）in 2003,

teristics of a universal adhesive in each of two modes:

manufacturers began exploring the usage of 10-MDP in

etch-and-rinse and self-etch. The two null hypotheses

combination with other components in novel adhesive

tested were as follows:（i）etching mode would make no

formulations. Universal adhesives that can be used withdifference to the dentin bond fatigue resistance, and（ii）

2）

resin cements on different substrates , with shortened

etching mode would make no difference to the energy

application times3）, or with different surface moisturecharacteristics of universal adhesive-treated dentin.

4）

conditions on enamel and dentin substrates have beenbrought to market. On the contrary, the bond durability

Materials and Methodsof universal adhesives has been reported to be inferior
to that of two-step self-etch adhesives5）. Nevertheless,

1 ．Study materials

the flexibility of universal adhesives has ensured their

The universal adhesive used in this study was Clear-

increasing popularity in clinics6）, creating a need to fur-fil Universal Bond Quick（CU, Kuraray Noritake Dental,

ther investigate better ways to utilize universal adhe-Tokyo, Japan）. A phosphoric acid-etching agent（Ultra-

sives.

Etch; Ultradent Products, South Jordan, UT, USA）and

Over the past decade, a method of assessing bonda composite resin（Z100 Universal Restorative; 3M Oralfatigue resistance in adhesives has been developed in a

Care, St. Paul, MN, USA）were employed to constructcollaborative effort between the Creighton University

the specimens（Table 1）.

School of Dentistry（CU, Omaha, NE, USA）and the

2 ．Specimen preparation

Nihon University School of Dentistry（NU, Tokyo,

De-identified extracted human molar teeth were used

Japan）. Development began with Erickson et al.7） andin this study. The experimental protocol for using

has been refined by researchers from CU and NU8‒10）.

de-identified human molar teeth was reviewed and

9）

10）

The appropriate frequency , number of cycles , andapproved by the Biomedical Institutional Review Board

method of analysis11） have been established. However,at the CU（No. 760765‒1）and the Ethics Committee forthe universal adhesive made by the manufacturer that

Human Studies of the NU（No. 2015‒06）.

developed 10-MDP has not been investigated with thisSectioned buccal and lingual halves of the teeth withestablished testing method, although such an investiga-

the apical portions removed were mounted in 25 mmtion could provide novel insights for the further devel-

brass rings using an acrylic resin（Bosworth Fastray;

opment of universal adhesives.

Keystone Industries, Myerstown, PA, USA）. Flat dentin

12）

Yoshida et al.

reported that the chemical interac-surfaces were prepared on the mounted buccal and lin-tion of 10-MDP with dentin is essential in obtaining

gual surfaces by wet-grinding using a sequence of sili-

showed that chemi-

con carbide（SiC）papers（Struers, Cleveland, OH, USA）

cal interactions between 10-MDP and dentin can beof gradually increasing fineness（#180-, #320-, #600-,

13）

durable bonds, while Inoue et al.

explained through changes in dentin surface character-#1,200-, #2,000-, and #4,000-grit）in a grinder-polisher

istics. Although it has been shown that the bonding（Ecomet 4; Buehler, Lake Bluff, IL, USA）. As the direc-

strategy employed does not significantly affect thetionality of surface scratches created by the abrasives

bonding performance of a given universal adhesive, it

might substantially influence the bond strength testingremains possible that the chemical bonding interactions

results, the surfaces were polished up to #4,000-grit toof a universal adhesive with ground dentin and withminimize this influence. These surfaces were thenetched dentin differ. Moreover, further assessment of

washed with water and dried using a dental three-way

changes in the energy characteristics of dentin surfaces

syringe at a distance of 5 cm above the surface at anDec, 2021

Dentin Bond Effectiveness of a Universal Adhesive
Table 1
Material（lot no.）ClearfilUniversal
Bond Quick（1L0003）

Type of material（code）

3

Materials used in this study
Main component（pH）

Manufacturer

bis-GMA, HEMA, 10-MDP,
hydrophilic amide monomer,
Universal adhesive
Kuraray Noritake Denethanol, water, initiators,（CU）tal, Tokyo, Japansilica, silane coupling agent（2.3）

Ultra-Etch（G019）

Etching agent

35％ phosphoric acid, glycol,
cobalt aluminate blue spinel

Ultradent Products,
South Jordan, UT, USA

Z100
Universal
Restrative（1312131）

Composite resin

bis-GMA, TEGDMA, silane3M Oral Care, St. Paul,treated ceramic, benzotriMN, USAazolyl methylphenol

bis-GMA: 2,2-bis［p（2-hydroxy-3-methacryloxy propoxy）phenyl］propane; HEMA:
2-hydroxyethyl methacrylate; 10-MDP: 10-methacryloyloxydecyl dihydrogen phosphate;
TEGDMA: triethylene glycol dimethacrylate

air pressure of 0.3 MPa.

A chisel-shaped metal rod was used to apply the load

Thirty specimens were prepared for each adhesiveto the metal ring（mold-enclosed method）immediately

for initial bond strength testing, and 40 specimens wereadjacent to the flat dentin surface. Fifteen specimens

prepared for each adhesive for bond fatigue strength

per group were loaded to failure using an all-electric

testing. Additionally, 20 specimens were prepared for

dynamic test instrument（ElectroPuls E1000; Instron,

each adhesive for surface free energy measurements.

Canton, MA, USA）at a crosshead speed of 1 mm/min.

Half of the specimens for each adhesive were phos-

Initial bond strength（MPa）was calculated from thephoric acid-etched for 15 s before application of thepeak load at failure divided by the bonded surface area.

adhesive（etch-and-rinse mode）, while the other half

4 ．Bond fatigue strength testing

were not etched（self-etch mode）. The specimens were

A staircase method was used to perform the bondprepared under ambient laboratory conditions of 23±

fatigue strength tests using the all-electric dynamic test

2℃ and 50±10％ relative humidity.

instrument.11） In the staircase method, the initial stress3 ．Initial bond strength testinglevel was set to half of the measured initial bond

Stainless steel（SUS304）rings with an inner diameter

strength and was applied to the first specimens. Theof 2.4 mm, an outer diameter of 4.8 mm, and a height offatigue load was applied to the metal ring using a sine

2.6 mm were used to bond a composite to the dentin.

wave at a frequency of 20 Hz for 50,000 cycles or until

The bonding side of each metal ring was treated with afailure occurred. If the first specimen survived the

3％ solution of paraffin in hexane. Each dentin surface

application of 50,000 cycles, a new second specimen was

was treated with the universal adhesive in etch-and-tested for the same number of cycles at a stress level

rinse or self-etch mode according to the manufacturer’sthat was about 10％ higher. If the first specimen failed,instructions. A custom fixture was used to position andthe stress level was reduced by the same amount for

hold the stainless steel ring over the bonding site as thethe second specimen. This procedure was repeated forcomposite was placed into the ring using a condensing

20 specimens, raising and lowering the stress based on

instrument. The composite was then light-cured for 40 s

survival or failure at each loading force. The test speci-

using a quartz-tungsten-halogen（QTH）curing unit

mens were immersed in water at room temperature（Spectrum 800 Curing Unit; Dentsply Caulk, Milford,（23±2℃）during the testing.

DE, USA）set at a light intensity of 800 mW/cm2. The

5 ．Surface free energy measurement

bonded specimens were then stored in 37℃ distilled

The dentin surfaces were prepared as describedwater for 24 h before testing.

above. Each dentin surface was treated with universal

ODEP Vol. 1, No. 1

4adhesive in etch-and-rinse or self-etch mode in accor-

facturer’s instructions in etch-and-rinse or self-etchdance with the manufacturer’s instructions, and themode. The resin composite was then placed and light-ir-

uncured adhesive layer was removed by three set ofradiated for 40 s using the QTH curing unit from aalternating rinses of acetone and distilled water. The

standardized distance of 1 mm. For the composite/den-contact angles of the specimens were then measured totin interfaces, bonded specimens were embedded inanalyze the surface free energy characteristics of each

epoxy resin（Epon 812; Nisshin EM, Tokyo, Japan）andadhesive-treated surface. Phosphoric acid-etched andthen stored at 37℃ for 24 h. Each bonded specimen

ground dentin surfaces were also measured. The sur-was then sectioned near its center, and the surfaces offace free energy characteristics of the specimens werethe cut halves were polished with a sequence of SiC

determined by measuring the contact angle formed

papers（#180-, #320-, #600-, #1,200-, #2,000-, and #4,000-

with the surface by the three test liquids of known sur-grit）using a grinder-polisher. Finally, the surface wasface free energy parameters: 1-bromonaphthalene,

polished with a soft cloth using 1.0μm particle diameter

diiodomethane, and distilled water. For each test liquid,

diamond paste（DP-Paste; Struers, Ballerup, Denmark）.

the equilibrium contact angle（θ）was measured using

SEM specimens of the composite-dentin interfaces were

the sessile drop method under ambient laboratory con-dehydrated by first immersing them in ascending con-ditions, as described earlier, using a contact angle mea-

centrations of aqueous tert-butanol（50％ for 20 min,surement apparatus（DM 500; Kyowa Interface Science,

75％ for 20 min, 95％ for 20 min, and 100％ for 2 h）and

Saitama, Japan）for ten specimens per group. The appa-

then transferring them from the final 100％ bath to aratus was fitted with a charge-coupled device camera to

freeze-drying apparatus（Model ID-3; Elionix）
for 30 min.

enable automatic measurement. A standardized 1.0μl

The polished surfaces were etched for 30 s using an

drop of each test liquid was placed on the treated den-argon ion beam（Type EIS-200ER; Elionix）directed

tin surface, and a profile image was captured after 500

perpendicular to the surface at an accelerating voltage

ms using the apparatus. The contact angle was thenof 1.0 kV and an ion current density of 0.4 mA/cm2.

calculated using the θ/2 method, using the built-in

The surfaces were then coated with a thin film of gold

interface measurement and analysis system（FAMAS;

in a vacuum evaporator（Quick Coater Type SC-701;

Kyowa Interface Science）. The surface free energy（γs）

Sanyu Electron, Tokyo, Japan）and observed using

and its parameters for solids（γsd, γsp, and γsh）were

field-emission SEM at an operating voltage of 10 kV.

calculated using the formulae described by Hata et
14）

7 ．Statistical analysis

al. , again using the built-in software（FAMAS）. The

Initial bond strength data were analyzed with one-dispersion force（γsd）represents the weakest intermo-

way analysis of variance（ANOVA）followed by Tukey’s

lecular force between apolar molecules. The polar

post hoc honestly significant difference（HSD）test. The（non-dispersion）force（γsp）represents the electric andbond fatigue strength data were analyzed using a modi-

metallic interactions, in addition to the dipolar interac-fied t-test with a Bonferroni correction and custom soft-

tions. Besides these two parameters of γs, the hydro-

ware. The γs, γsd, γsp, and γsh data were analyzed using

gen-bonding force（γ ）, which relates to the water andone-way ANOVA along with Tukey’s HSD test. All

hydroxyl components, was calculated.

statistical analyses, apart from the bond fatiguesh

6 ．Scanning electron microscopy obser vation ofbonding interface
Representative scanning electron microscopy（SEM）
micrographs of the resin-dentin interfaces for threestrength data analysis, were conducted using statisticalsoftware（SPSS Statistics Ver. 13; International Business
Machines, Armonk, NY, USA）, applying a significancelevel of 0.05.

specimens per group were obtained using field-emission
SEM（ERA 8800FE; Elionix, Tokyo, Japan）. A rectangu-

Resultslar（4×2×1 mm）section of dentin was removed fromeach molar for SEM observations of the bonding inter-

The initial bond strength of the universal adhesive in

face. The dentin surface was prepared as describedeach etching mode is shown in Table 2. The initial bondabove for specimen preparation. The dentin surface

strength of the universal adhesive was not significantly

was treated with the adhesive according to the manu-

influenced by the etching mode（p＝0.606）.

Dec, 2021
Table 2

Dentin Bond Effectiveness of a Universal Adhesive

Initial bond strength of universal adhesives to den-Table 3tin using etch-and-rinse and self-etch modes
Code

Etch-and-rinse mode

CU

Code

a

27.3（4.2）

CodeEtch-and-rinse modeaCU

28.1（4.2）

Unit: MPa. Values in parentheses indicate standard deviations. Same small letter indicates no significant difference（p＞0.05）.

Table 4

Bond fatigue strength of universal adhesives todentin using etch-and-rinse and self-etch modes

Self-etch modea5

13.9（1.9）

Self-etch mode
14.4（1.7）a

Unit: MPa. Values in parentheses indicate standard deviations. Same small letter indicates no significant difference（p＞0.05）.

Surface free energy characteristics of universal adhesive-treateddentin using etch-and-rinse and self-etch modes
γs

Etch-and-rinse mode
γsd

γsp

γsh

γs

Self-etch mode
γsd

γsp

γsh

baseline

37.8
1.1
2.2
68.8
41.0
2.8
25.0
41.1
a
a
a
a
a
（1.5）
（1.6）a （1.1）a （3.6）（1.4）（2.3）a （2.4）
（2.5）

CU

39.2
37.2
1.1
0.9
64.1
40.2
2.5
21.4
a
a
a
（2.4）
（0.8）
（1.8）a （2.1）b （2.9）b （1.1）（1.5）a （2.0）b

Unit: mN/m. γs, surface free energy; γsd dispersion force; γsh, hydrogen-bonding force; γsp, polar force. The same letters in a column indicate no statistically significant difference（p＞0.05）.

The bond fatigue strength for the universal adhesive

tubules.

in each etching mode is shown in Table 3. The bondfatigue strength of the universal adhesive was not sig-

Discussionnificantly influenced by the etching mode.
The surface free energy parameters of the universal

In the present study, etching mode did not affect theadhesive-treated dentin are shown in Table 4. The

dentin bond fatigue resistance of the universal adhesive,

baseline in etch-and-rinse mode exhibited significantly

and thus the first null hypothesis was not rejected. A

lower γs and γsh values than in self-etch mode（p＜0.001

recent study15）, which used the same research design tofor γs and γsh）. Changes in the γsd and γsp values of thestudy enamel bonding, reported that the bond fatigue

universal adhesive-treated dentin were not significantly

resistance of universal adhesives was significantlysddecreased（γ : p＝0.279 for etch-and-rinse mode, p＝higher in etch-and-rinse mode than in self-etch mode.

0.571 for self-etch mode; γsp: p＝1.000 for etch-and-rinse

The results of the present and previous studies suggest

mode, p＝0.734 for self-etch mode）by the adhesivethat the use of etch-and-rinse or selective etching

application, in contrast to γs and γsh in self-etch mode,

modes with universal adhesives is more effective thanwhose values were significantly decreased（γs: p＝0.004;

that of self-etch mode from the viewpoint of bond

γsh: p＝0.002）.

fatigue resistance, in agreement with a systematic

Representative SEM images of the composite-dentin
interfaces of etch-and-rinse samples and self-etch sam-review of earlier laboratory bond strength evaluations16）.

ples are shown in Fig. 1. The thickness of the layer of

The results for interfacial characteristics showed that

universal adhesive was ‒10μm. For each tested adhe-the baseline γs , γsd , γsp , and γsh values for dentin in

sive, the composite-dentin interface showed excellent

etch-and-rinse mode were significantly lower than in

adaptation to dentin regardless of etching mode. How-

self-etch mode. These results indicate that the wettabil-

ever, deeper penetration of adhesives into the dentinal

ity, hydrophilicity, and degree of polarization of phos-

tubules was observed in etch-and-rinse mode due tophoric acid-etched dentin might be lower than those ofremoval of the smear layer and opening of the dentinal

ground dentin. Tay et al.17） reported that the dehydra-

ODEP Vol. 1, No. 1

6

Fig. 1 Scanning electron microscopy images of the composite-dentin interface
One panel as CU etch & rinse and the other panel as CU self-etch at（a）2,500×and（b）10,000×
magnifications. A, adhesive; D, dentin; HL, hybrid layer; R, resin composite.

tion of demineralized dentin resulted in osmosis oflayered structure of 10-MDP and calcium salt. A

water content from deeper dentin, leading to weakergreater extent of chemical bonding to the dentin sur-bonding due to osmotic blisters and hydrolysis of theface should therefore create a more hydrophobic sur-adhesive itself. However, etching mode did not influ-

face. Only the γsh value of adhesive-treated dentin inence the dentin bond fatigue resistance of the universal

etch-and-rinse mode was significantly lower than theadhesive in the present study. In our SEM observationsb a s e l i n e , a l t h o u g h b o t h γs a n d γs h v a l u e s w e r eof adhesive interfaces, deeper penetration of the adhe-decreased after adhesive application in self-etch mode.

sive into the dentinal tubules was observed in etch-and-Thus, the second null hypothesis was rejected. A previ-

rinse mode due to removal of the smear layer andous study13） reported that interfacial characteristics

opening of the dentinal tubules by phosphoric acid etch-were influenced by reactions between acidic functionaling, despite the lower wettability of the dentin. Thesemonomers and calcium in tooth substrates; thus thedeeper resin tags, lying perpendicular to shear stress,

present results are consistent with those of previous

may have contributed somewhat to the cyclic fatigue

studies. Based on the different tendencies observed inresistance of adhesive interfaces. Additionally, bondthe changes in interfacial characteristics between base-fatigue testing in this study was performed with a fre-line and adhesive-treated dentin, the chemical interac-quency of 20 Hz and a cycling period of 50,000 cycles,

tions in self-etch mode might be much stronger thanbut only for ‒40 min, which may not be a sufficiently

those in etch-and rinse mode.

long testing period. Hence, the influence of osmosis ofAnother interpretation of the results of bond fatigue

water content from deeper dentin, resulting in osmotic

testing might be supported by these surface freeblisters and hydrolysis of adhesive, might have beenenergy changes. Even with the deeper penetration of

reduced due to the short testing period compared toresin tags, lower reactivity between 10-MDP and colla-

tests using long-term water storage or thermal cycling.

gen fibrils might reduce the stability of the bond, add-

For the universal adhesive employed in this study,

ing to the effects of the lower wettability of the adher-

10-MDP is a key technological factor for chemical bond-end surface. However, further research is needed toing with dentin substrate, regardless of the bonding

clarify the contribution to dentin bond durability of thestrategy employed. The adhesion-decalcification con-interaction between the functional resin monomer and

cept

18）

claimed that the functional group of 10-MDP

naked collagen.

interacts ionically with calcium in dentin, forming a

The clinical implication of this study is that althoughchemically bonded structure of 10-MDP and calcium

many Japanese clinicians use universal adhesive insalt layers on the dentin surface. As the long carbonyl

selective etching mode, there is not much need to19）

chain of 10-MDP is relatively hydrophobic , a hydro-

worry about exposure of dentin to phosphoric acidphobic layer would cover the dentin surface due to thewhen the clinician places a direct composite filling with

Dec, 2021

Dentin Bond Effectiveness of a Universal Adhesive

this adhesive.

74）Kumagai RY, Hirata R, Pereira PNR, Reis AF. Moist vsConclusion
1．The initial bond strengths and bond fatigue
strengths were not significantly influenced by etchingmode.
2．The baseline in etch-and-rinse mode exhibited significantly lower γs and γs h values than in self-etchmode. Only the γsh value of adhesive-treated dentin in
etch-and-rinse mode was significantly lower than theb a s e l i n e , w h i l e b o t h t h e γs a n d γs h v a l u e s w e r edecreased after adhesive application in self-etch mode.
3．Based on SEM observations, the adaptation of theinterface to dentin was excellent for both etchingmodes.
Conflicts of interest
The authors of this manuscript certify that they have no
proprietary, financial, or other personal interest of any
nature in terms of any product, service, and/or company
that is presented in this article.
Acknowledgments
This work was supported in part by Grants-in-Aid for Scientific Research（grant no. 19K10158, 21K09900）from theJapan Society for the Promotion of Science. This project wasalso supported in part by the Sato Fund, by the Uemura
Fund, and by a grant from the Dental Research Center of
the Nihon University School of Dentistry, Japan. The
authors thank Mr. Jason M. Moody of the Creighton University School of Dentistry for technical contributions.

References
1）Van Meerbeek B, Yoshihara K, Van Landuyt K, Yoshida
Y, Peumans M. From Buonocore’s pioneering acid-etch
technique to self-adhering restoratives. A status perspective of rapidly advancing dental adhesive technology. J Adhes Dent 2020; 22: 7‒34.
2）Cuevas-Suárez CE, de Oliveira da Rosa WL, Vitti RP, daSilva AF, Piva E. Bonding strength of universal adhesives to indirect substrates: A meta-analysis of in vitro
studies. J Prosthodont 2020; 29: 298‒308.
3）Saikaew P, Matsumoto M, Chowdhury AFMA, Carvalho
RM, Sano H. Does shortened application time affectlong-term bond strength of universal adhesives to dentin?. Oper Dent 2018; 43: 549‒558.

over-dried etched dentin: FE-SEM/TEM and bondstrength evaluation of resin-dentin interfaces produced
by universal adhesives. J Esthet Restor Dent 2020; 32:
325‒332.
5）Tsujimoto A, Barkmeier WW, Takamizawa T, Watanabe H, Johnson WW, Latta MA, Miyazaki M. Comparison between universal adhesives and two-step self-etchadhesives in terms of dentin bond fatigue durability in
self-etch mode. Eur J Oral Sci 2017; 125: 215‒222.
6）Nagarkar S, Theis-Mahon N, Perdigão J. Universal dental adhesives: Current status, laboratory testing, andclinical performance. J Biomed Mater Res B Appl Biomater 2019; 107: 2121‒2131.
7）Erickson RL, De Gee AJ, Feilzer AJ. Effect of pre-etching enamel on fatigue of self-etch adhesive bonds. Dent
Mater 2008; 24: 117‒123.
8）Barkmeier WW, Erickson RL, Latta MA. Fatigue limitsof enamel bonds with moist and dry techniques. Dent
Mater 2009; 25: 1527‒1531.
9）Scheidel DD, Takamizawa T, Bakmeier WW, Erickson
RL, Tsujimoto A, Miyazaki M. Effect of frequency on
the fatigue strength of dentin bonds. J Oral Sci 2016; 58:539‒546.
10）Tsujimoto A, Barkmeier WW, Erickson RL, Takamizawa T, Latta MA, Miyazaki M. Influence of thenumber of cycles on shear fatigue strength of resin
composite bonded to enamel and dentin using dental
adhesives in self-etching mode. Dent Mater J 2018; 37:
113‒121.
11）Tsujimoto A, Barkmeier WW, Erickson RL, Fischer NG,
Markham MD, Takamizawa T, Latta MA, Miyazaki M.
Shear fatigue strength of resin composite bonded todentin at physiological frequency. Eur J Oral Sci 2018;
126: 316‒325.
12）Yoshida Y, Yoshihara K, Nagaoka N, Hayakawa S, Torii
Y, Ogawa T, Osaka A, Van Meerbeek B. Self-assembled
nano-layering at the adhesive interface. J Dent Res
2012; 91: 376‒381.
13）Inoue N, Tsujimoto A, Takimoto M, Ootsuka E, Endo H,
Takamizawa T, Miyazaki M. Surface free-energy measurements as indicators of the bonding characteristics
of single-step self-etching adhesives. Eur J Oral Sci
2010; 118: 525‒530.
14）Hata T, Kitazaki Y, Saito T. Estimation of the surface
energy of polymer solids. J Adhes 1987; 21: 177‒194.
15）Nagura Y, Tsujimoto A, Barkmeier WW, Watanabe H,
Johnson WW, Takamizawa T, Latta MA, Miyazaki M.
Relationship between bond fatigue durability and surface free energy characteristics of universal adhesives
to enamel. Eur J Oral Sci 2018; 126: 135‒145.
16）Rosa WL, Piva E, Silva AF. Bond strength of universal

ODEP Vol. 1, No. 1

8adhesives: a systematic review and meta-analysis. J

beek B. Adhesion/decalcification mechanisms of acid

Dent 2015; 43: 765‒776.
17）Tay FR, Pashley DH, García-Godoy F, Yiu CK. Sin-interactions with human hard tissues. J Biomed Mater
Res 2002: 59: 56‒62.

gle-step, self-etch adhesives behave as permeable membranes after polymerization. Part Ⅱ. Silver tracer penetration evidence. Am J Dent 2004; 17: 315‒322.
18）Yoshioka M, Yoshida Y, Inoue S, Lambrechts P, Vanherle G, Nomura Y, Okazaki M, Shintani H, Van Meer-19）Van Landuyt KL, Yoshida Y, Hirata I, Snauwaert J, De
Munck J, Okazaki M, Suzuki K, Lambrechts P, Van
Meerbeek B. Influence of the chemical structure offunctional monomers on their adhesive performance. J
Dent Res 2008; 87: 757‒761.

Dec, 2021

9

Original Article

ODEP 1（1）：9～21，2021

Development of Chairside Dry Heat Sterilizer Using Halogen Lamps
as a Thermal Radiation Source
Keisuke SAIGUSA, Yukihiro TAKAHASHI＊,
Munehiro MAEDA and Masaru IGARASHI
Department of Endodontics, The Nippon Dental University, School of Life Dentistry at Tokyo＊

Department of Microbiology, The Nippon Dental University, School of Life Dentistry at Tokyo

Abstract
Purpose: A chairside dry heat sterilizer using halogen lamps as a heat radiation source was developed, andits potential for clinical application was investigated.
Methods: The temperature in the sterilization chamber was measured in two groups: one group in which
the central temperature was set and maintained at 330℃, and another group in which the central temperature continuously increased. Then, changes in the physical properties of a stainless-steel K-file irradiated withthe lamp were examined by observing the surface by scanning electron microscope and by bending and torsion tests. Finally, sterilization tests were conducted using spore-forming bacteria and indigenous oral bacteria.
Results: The temperature in the light condensing area stabilized after about 20 s, with little effect on thephysical properties of the K-file. Sterilization of spore-forming bacteria took several tens of seconds, whilethat of indigenous oral bacteria took 3‒5 s on a paper point and 7‒10 s on a cotton ball.
Conclusion: It is suggested that the chairside dry heat sterilization using a thermal radiation source is useful for dental materials.
Key words: simple dry heat sterilizer, halogen lamp, stainless-steel K-file, strength test, spore-forming bacteria

Corresponding author: Keisuke SAIGUSA, Department of Endodontics, The Nippon Dental University, School of Life Dentistryat Tokyo, 1‒9‒20, Fujimi, Chiyoda-ku, Tokyo 102‒8159, Japan
TEL: ＋81‒3‒3261‒5698, FAX: ＋81‒3‒5216‒3718, E-mail: k-saigusa_d2117007@tky.ndu.ac.jp
Received for Publication: January 6, 2021/Accepted for Publication: January 15, 2021
DOI: 10.11471/odep.2021‒002

ODEP Vol. 1, No. 1

10

to the sterilized instrument and be transported directly

Introduction

into the root canal, resulting in blockage of thecanal11,12）, harm to the human body due to heavy metal

Aseptic treatment in dentistry is one of the most

poisoning, and burns due to scattering of hot material.

important principles to improve treatment outcomes

Meanwhile, other sterilizers also have problems, suchand prevent the spread of infection via treatment

as unbalancing the table of the dental unit due to the

instruments. In particular, endodontic treatment

excessive weight of the entire sterilizer, media spillage

requires thorough removal of bacterially infected toothwhen the sterilizer is tipped over, and salt coagulation

structures and antiseptic treatment of healthy dentin.

due to humidity in hot salt sterilizers. Furthermore, all

In addition, root canal treatment possesses the risk ofsterilizers take time to heat the media, so advance

contamination by blood and saliva during the proce-

preparation is necessary. Finally, in recent years, oppor-dure, and also by necrotic tissue that has been infected

tunities for home-visit dental care, in addition to dental

by bacteria, making it necessary to re-disinfect thecare in the clinic, have been increasing, and thus there

treatment instruments during the procedure.

is a demand for easy, safe, and portable sterilizers that

In recent years, from the perspective of nosocomial

can be brought into a typical residence.

infection, dental care workers have needed to be espe-

In this study, we developed a simple dry heat steril-cially careful about their own infection potential to pre-izer that uses halogen lamps as a thermal radiation

vent bacterial infections caused by methicillin-resistantsource to convert electrical energy into thermal energy

Staphylococcus aureus, Pseudomonas aeruginosa, andand directly apply it to materials to be sterilized in the

Treponema pallidum

1,2）. Viral infections include infec-

form of electromagnetic waves. Then, we investigatedtions caused by viruses of the herpesvirus family, hepa-

the characteristics of the sterilizer to determine its

titis B and C viruses, and human immunodeficiency

potential for clinical application.

virus, and these microorganisms have also been
detected in dental pulp and periodontal tissues 3‒5）.

Materials and Methods

Recently, severe acute respiratory syndrome coronavirus 2（SARS-CoV-2）has caused a worldwide pandemic
and the infection is still spreading, so patients must betreated with caution during dental care6,7）.

1 ．Experimental equipment
An overview of the prototype sterilizer and peripheral apparatus is shown in Fig. 1. The prototype steril-

Small sharp instruments such as reamers and files,

izer was composed of two metal plates, each 154 mm inwhich are frequently used in endodontics, are classified

width by 70 mm in height, and two heat sinks, eachas critical instruments and are sterilized by gas or

100 mm in width by 70 mm in height. These were com-high-pressure steam sterilization before use8）. However,bined on a metal chassis measuring 180 mm wide byonce they come into contact with fingers, saliva, or

120 mm deep by 55 mm tall. A hole 1 cm in diameter

infected dentin, bacterial contamination of the instru-

was made in the center of the upper metal cover, andments is inevitable, and because these instruments arehalogen lamps were installed in the heat sinks.

used repeatedly during the procedure, frequent sterilization during treatment is necessary.

A halogen lamp spot heater（HSH-30, Fintech, Hyogo,
Japan）with a focal length of 40 mm, a light condensing

Many endodontic dry heat sterilizers that can bearea diameter of 10 mm, and power of 75 watts wasused chairside have been described in the literature,used for the experiment. Irradiation was carried out inincluding 1）devices using molten tin-based alloys, 2）both directions on opposite sides of the device’s coaxial

devices using salt as a medium, 3）bead sterilizers using

axis, and the material to be sterilized was placed in thesmall glass beads or metal balls, 4）electric furnace-typeirradiation field in the middle of the heater. The voltage

dry heat sterilizers with heaters wrapped aroundwas supplied by a 100 V household power supply, whichquartz tubes, and 5）sterilizers that heat brass blocks

was converted to 24 V for use, and the current was set

with slits 9‒18）. However, sterilizers that use molten

at 3.125 A.

metal or small balls, such as glass beads, introduce the

In Fig. 2, K-type thermocouples（Φ 0.3 mm）were

possibility that the metal or balls will become attachedplaced alternately facing each other at 5-mm intervals

Dec, 2021

Development of a Simple Dry Heat Sterilizer Using Halogen Lamp

11
100 mm

154 mmE

70 mm

Heat sink

55 mm

D

A

※

C

B

180 mm

120 mmLong side

Short sideabFig. 1（a）Overview of prototype sterilizer and peripheral apparatus：（A）Prototype sterilizer,（B）Control apparatus,（C）Power
supply unit,（D）Irradiation switch,（E）Temperature control apparatus（b）Schematic diagram of prototype sterilizer（※：Base of the halogen lamp）

40 mm

B

A

A

B

C

10 mm

C

a

b

Fig. 2（a）Prototype sterilizer（Fig. 1, A）, top view：（A）Halogen lamps,（B）K-type thermocouples,（C）Heatsinks（b）Cross-sectional schematic diagram of the prototype sterilizer：the entire circle is the light condensing area, and the circumference is the outer edge. The black points represent the tips of theK-type thermocouples（depths of 10, 15, 20, 25, 30, and 35 mm from the top）.

up to 35 mm from the side of the apparatus to measure

Ⅰ: When the control switch is turned on, the lamp

the temperature of the sterilization area. The light con-cycles on and off to reach and maintain the set tem-densing area（10 mm）at the focus of the halogen lamp

perature automatically.

was set at a depth of 15‒25 mm from the hole, with 15

Ⅱ: When the control switch is turned off, continuousand 25 mm at the outer edge of the light condensing

irradiation is performed and the temperature increases

area and 20 mm at the center of the light condensing

until the switch is turned off.

area.

2 ．Experimental details

The irradiation time could be set from 0.1 to 99.9 s at

The following items were investigated to determine

0.1-s intervals, and a countdown system was adopted.

the effects of this device on endodontic instruments andThere was also a temperature control switch that could

materials.

be set from 100℃ to 500℃ in 1℃ increments. Two settings for halogen lamp irradiation were established:ODEP Vol. 1, No. 1

12
Load cell

Load cell
45°

3 mm

Fig. 3 Bending test machine
The first 3 mm of the file tip
is fixed with a chuck, and thebending torque is measured
when the file is bent to 45°.

Fig. 4 Torsion test machine
The first 3 mm of the file tip is fixed with achuck, the file is twisted until it breaks, and thetorsion torque and rotation angle at the time of
failure are measured.

the top, then the K-file was allowed to cool by natural
air cooling to room temperature, and this process was

1 ）Measurement of temperature in the light condensing area
The K-type thermocouples placed inside the sterilizer

repeated 10 times. We observed the surface changes by
SEM for each group（n＝1）.
3 ）Strength testing of files after heat irradiation

were used to record the temperature change over time

The changes in physical properties of the files after

at 100-ms intervals when the temperature was con-irradiation with halogen lamps were tested according totrolled at 330℃ with setting Ⅰ and when the tempera-

the international standard ISO 3630-119）for small dental

ture was continuously increased with setting Ⅱ. The

instruments and judged according to the standard val-

irradiation time was set to 90 s for both settings.

ues of ISO 3630-5.20）Two types of #15‒40 stainless-steel

The measurement data were recorded on a PC

hand K-file were used for the test: K-file and Senseus（Inspiron 15 7000 Gaming, DELL, Round Rock, TX,

K-FlexoFile（Dentsply Sirona, Charlotte, NC, USA）.

USA）using a data logger（NR-1000, KEYENCE, Osaka,

Since there was a risk of bias with a single type of sam-Japan）. Temperature measurements were performedple, we made a comprehensive judgment by comparing

at room temperature（25℃）and humidity（55％）.

two K-files with their respective untreated groups.

2 ）Surface observation of K-files after heat irradiation

Non-irradiated files were used as a control group, andthe experimental group consisted of files that were

The surface of #40 stainless-steel hand K-files（MANI,

irradiated 10 times with halogen lamps for 10 s at set-

Tochigi, Japan）
was observed under a scanning electronting Ⅱ（n＝10 for each group）. All instruments weremicroscope（SEM, S-4000, Hitachi, Tokyo, Japan）after

allowed to stand at room temperature（25℃）for 30 min

halogen lamp irradiation to detect any change in theafter each irradiation before the next irradiation. Before

surface properties.

each test, the handle of the file was removed with wire

Five experimental groups were established. The first

cutters at the point where it was attached to thegroup was subjected to natural air cooling to room tem-

instrument shaft.

perature after 5 s of irradiation, and this cycle was（1）Bending test（ISO 3630-1）

repeated 10 times at setting Ⅱ. The second and third

The bending test machine and test method aregroups were subjected to the same treatment, but atshown in Fig. 3. A section of the K-file up to 3 mm from

irradiation times of 10 s and 15 s. In addition, two con-the tip was fixed vertically in the chuck section of thetrol groups were set. An untreated group was used as abending test machine and rotated at a speed of 100

negative control and a flame-treated group was used as

mm/min to induce bending. The maximum bending

a positive control. In the flame-treated group, the blade

torque was measured and the average value of 10 files

was set at the position of the oxidizing flame of the gas

was used as the maximum bending torque（mN・m）.

burner（about 1,500℃）and heated for 10 s by graspingDec, 2021

Development of a Simple Dry Heat Sterilizer Using Halogen Lamp

13

－Sterility test－
Halogenlamp
K－fileHalogenlamp

K－filePaper point Cotton ball

Paper point

Cotton ball

3. Irradiate contaminatedmaterials with halogen lamps

1. Prepare solution ofeach bacteria species 2. Contaminate samples
with each solution

Fig. 5

4. After irradiation, placesamples in sterile Tryptic
soy broth

5. Incubate cultures at 37℃
for up to 2 wk and visually
determine the results

Sterilization test procedure（2）Torsion test（ISO 3630-1）

method, the spores were collected by centrifugation at

The torsion test machine and test method used are

3,000×g for 15 min, washed three times with sterile

shown in Fig. 4. Each sample was set in the chuck of

distilled water, and then heat-treated at 75℃ for 30 min

the motor, and a section of the file up to 3 mm from theto kill the vegetative form bacteria. Other bacteria

tip was fixed in the chuck on the opposite side. The

were inoculated in brain heart infusion（BHI: Becton,

torque measuring device on the top of the machine was

Dickinson and Company, Sparks, MD, USA）broth andconnected to the torsion testing machine by a wire, andincubated at 37℃ for 24 h.

the motor was operated at 100 mm/min. The average

For experimental samples, we used a #40 stain-

value of 10 files was used as the maximum torsional

less-steel hand K-file, #40 sterile paper points（Absor-

torque（mN・m）and fracture angle（°）.

bent Paper Points, Zipperer, Germany）, and cotton balls

4 ）Heat sterilization tests for K-files, paper points,
and cotton ballswith a diameter of 3 mm（Micro Cotton Ball #3, Iwatsuki, Tokyo, Japan）. The K-file was cut with wire nip-

Various microorganisms were applied to different

pers just below the handle, and the cotton balls weredental materials to evaluate the sterilization effective-

removed from their container, placed in sterilization

ness. Bacillus atrophaeus ATCC9372 was used because

bags, and sterilized by high-pressure steam in an auto-

it is regarded as an indicator organism for dry heatclave for the experiments.

21）

sterilization in ISO 11138-4 . Fusobacterium nucleatum

The procedure of the sterilization test is shown in

ATCC23726, Streptococcus intermedius NCTC11324,

Fig. 5. The K-file was immersed up to 16 mm from theActinomyces oris（formerly known as A. viscosus）

tip of the blade in a bacterial solution adjusted to 1.0×

T14V22）, and Streptococcus mutans MT814823） were

108 colony forming units（CFU）/ml for each species.

used as representative of indigenous oral bacteria. In

Contaminating bacteria on the paper points and cotton

addition, Escherichia coli W3110 was also included, for aballs were prepared at approximately 2.0×103 CFU/ml.

total of six bacteria species used in the experiment.

The paper points were immersed in 2.0×106 CFU/ml

B. atrophaeus and F. nucleatum were purchasedof bacterial solution up to 10 mm from the tip（aboutfrom the American Type Culture Collection（ATCC）,

1μl）, and the cotton balls were contaminated by drop-

and S. intermedius was purchased from the National

ping 10μl of 2.0×105 CFU/ml bacterial solution. The

Collection of Type Cultures（NCTC）. E. coli W3110 was

cotton balls were then placed in a sterile Petri dish for

obtained from the Institute of Medical Science, Univer-

30 min at 37℃ to dry them before use in the experi-sity of Tokyo.

ments.

First, only B. atrophaeus was inoculated in 50 ml of

For the K-files, the irradiation time was set to 0［Con-Shaeffer’s medium24） as a spore-forming medium, andtrol（＋）］, 3, 5, 7, and 10 s with setting Ⅱ for all bacte-cultured at 37℃ with shaking for 24 h. The sporesria. For the paper and cotton samples, setting Ⅰ waswere then removed from the shaking incubator andused, and the irradiation time was set to 0［Control

stored at room temperature for 1 wk. After confirming（＋）］, 3, 5, 7, 10, 15, 20, 30, and 40 s for B. atrophaeus

sufficient spore formation by Wirtz’s spore staining

and 0, 3, 5, 7, and 10 s for other bacteria（n＝20 for eachODEP Vol. 1, No. 1

14
500

500

450

450
400

20 mm

350
300

25 mm

250

15 mm

200

10 mm

150

30 mm

100

0

15 mm

300
250

10 mm

200

30 mm

150

35 mm

50

10 20 30 40 50 60 70 80 90 100 110 120130140 150160 170180 190 200
Time
（s）

10mm

25 mm

350

100

35 mm

50
0

Temperature（℃）

Temperature（℃）

400

20 mm

15mm

20mm

25mm

0

0

10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200
Time
（s）

30mm

35mm

Temperature measurement during irradiation
at setting Ⅰ
Irradiation was started 10 s after the start of tem-

10mm

15mm

25mm

30mm

35mm

Temperature measurement during irradiation
at setting Ⅱ
Irradiation was started 10 s after the start of tem-

Fig. 6

Fig. 7

perature measurement.

perature measurement.

group）. The test samples were irradiated with halogen

20mm

until the end of irradiation.

lamps and incubated in sterile Tryptic soy broth（Bec-

At 15 mm and 25 mm, which were the outer edges ofton, Dickinson and Company）at 37℃ for 24 h, 48 h, 72

the condensing area, the temperature rose to approxi-

h, 1 wk, and 2 wk, the results were visually checked,

mately 260℃ and remained at around 240‒250℃ until

and those with increased turbidity or precipitation

the end of irradiation. The temperature dropped

were judged as positive for culture. For each group, thesharply outside the condensing area, reaching only 120‒number of samples that were culture negative was

150℃ at 10 mm and 30 mm and 70‒90℃ at 35 mm.

counted, divided by 20（the total number of samples）,

In setting Ⅱ, continuously increasing irradiation wasand multiplied by 100 to calculate the culture negative

performed for 90 s, and the results were measured overachievement rate.

time（Fig. 7）. In the light condensing area at a depth of

The paper points were cut 10 mm from the contaminated tip before being fed into the sterilizer.

15‒25 mm, the temperature rose rapidly as soon as
irradiation started, reaching 275℃ at the center and3 ．Statistical analysis of strength215‒226℃ at the outer edge 5 s after the start of irradi-

Student’s t-test was performed on the data obtained

ation. After another 10 s, the temperature reachedfrom the strength tests. The statistical significance

368℃ at the center and 280‒289℃ at the outer edge,

level was set at less than 5％, and IBM SPSS ver. 25

and after 15 s, 408℃ at the center and 307‒317℃ at thewas used as the statistical software.

outer edge. The highest temperatures after 90 s of irradiation were 464℃ at the center and 367℃ at the outer

Resultsedge.
2 ．Surface observation of K-files after heat irradi-

1 ．Temperature measurement of sterilization
area

ation
The surface of each file was observed by SEM（Fig.

The temperature of the sterilization area was mea-

8）. The flame-treated group showed a loss of continuity

sured for 90 s when the temperature was controlled todue to the collapse of the file edge, with the metal onaround 330℃ with setting Ⅰ（Fig. 6）. When this tem-the file surface peeling off. In the group irradiated byperature was reached, the control apparatus could not

the halogen lamp, there was no major change in thereact quickly enough, and the center temperature rose

surface properties of the files irradiated for 5, 10, andto 349℃. However, 20 s after the start of control, the15 s compared with the untreated group, either bytemperature stabilized at around 330℃. The amplitudevisual inspection without magnification or in the SEM

during each on/off switching cycle was about 10℃,

images.

which maintained the temperature at around 325‒335℃

Dec, 2021

Development of a Simple Dry Heat Sterilizer Using Halogen Lamp

15

Flame treatment group［Control（+）］

Untreated group［Control（－）］

5 s irradiation repeated 10 times group10 s irradiation repeated 10 times group15 s irradiation repeated 10 times group

Fig. 8

Surface observation by SEM of K-file irradiated by halogen lamp

3 ．Strength testing of K-files after heat irradiation

K-FlexoFile. The bending torque values in both groups

The results of the strength tests are shown in Fig. 9.

were within the ISO standard values.

1）Bending test2）Torsion test

The torques required to bend the files up to 45°are

The maximum torque and fracture angle until frac-shown in Fig. 9A and B. There was a significant differ-ture when torsion was applied to the file are shown inence between the control and irradiated groups for the

Fig. 9C‒F. For the #25 MANI K-file, results for the irra-#15 MANI K-files（p＜0.05）, and there was no signifi-

diated group were significantly lower than those for thecant difference between the groups for the Senseus

untreated group, and for the #35 MANI K-file, results

ODEP Vol. 1, No. 1
Bending torque（mN・m）

25
20
15
10
＊

5
0

#15

#20

#25

#30

Torsional torque（mN・m）

Control

#40

A
25
20
15

＊

10

＊

5
0

#15

#20

#25

#30

#35

#40

1,000
500
#20

#25

Control

#30

10
5
0

#15

#35

#40

Heat

E

#20

#25

#30

#35

#40

#35

#40

#35

#40

Heat

B
25
20
15
10
5
0

#15

#20

#25

ControlAngular deflection（ ° ）

1,500

#15

15

*p<0.05

Heat

2,000

0

20

Control

C

2,500

25

*p<0.05

Heat

ControlAngular deflection（ ° ）#35Torsional torque（mN・m）

Bending torque（mN・m）

16

#30
Heat

D

2,500
2,000
1,500
1,000
500
0

#15

#20

#25

Control

#30
Heat

F

Fig. 9 Bending test and torsion test results（p＜0.05, n＝20）（A）MANI K-file bending test results；（B）Senseus K-FlexoFile bending test results；（C）（E）MANI,K-file torsion test results；（D）（F）Senseus,
K-FlexoFile torsion test results

for the untreated group were significantly lower thanThe culture results for cotton balls were all negative

those for the irradiated group（p＜0.05）. The torqueafter 40 s for B. atrophaeus, after 10 s for S. mutans, A.

failure values for both groups were within the ISO

oris, and F. nucleatum, and after 7 s for other bacteria.

standard values.
There was no significant difference between the

Discussion

groups for Senseus K-FlexoFile, but only one of the 10#40 K-files had a fracture angle below the ISO standard

Most conventional simple dry heat sterilizers provideof 360°. Therefore, 10 additional tests were conducted

sterilization heat through a heating medium, such asin accordance with the standard, and all of them wereglass pellets or molten metal, and it takes several min-confirmed to meet the standard.

utes for the heating medium to rise to the sterilization

4 ．Sterilization tests

temperature. In contrast, the prototype of the simple

The results of the sterilization tests are shown in

dry heat sterilizer used in this study heats the materi-

Tables 1‒3. The culture results for K-files were all neg-als to be sterilized by directly radiated heat, which has

ative after 10 s of irradiation for B. atrophaeus and after

the advantage of eliminating the need to turn on the

7 s for other bacteria. Results for the paper points werepower supply and heat the medium beforehand because

negative after 30 s for B. atrophaeus, after 5 s for S.

there is no heating medium. Therefore, it is possible tomutans and A. oris, and after 3 s for other bacteria.

rapidly heat an object by turning on the power andDec, 2021

Development of a Simple Dry Heat Sterilizer Using Halogen Lamp
Table 1

Species and Irradiation

17

Sterilization test results of K-file（％）

24 h

48 h

72 h

1 wk

2 wk

0s

0

0

0

0

0

0s

0

0

0

0

0

3s

0
70

0
0

0
0

0
0

0
0

3s
5s

60
65

60
65

60
65

60
65

60
65

80

35

30

30

30

7s

100

100

100

100

100

10 s

100

100

100

100

100

10 s

100

100

100

100

100

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

B. atrophaeus

5s
7s

S. mutans

0s
3sSpecies and Irradiation
E. coliS. intermedius

24 h

48 h

72 h

1 wk

2 wk

0

0

0

0

0

0s

0

0

0

0

0

90

80

80

80

80

3s

80

80

80

80

80

5s

95

95

95

95

95

5s

95

95

95

95

95

7s

100

100

100

100

100

7s

100

100

100

100

100

10 s

100

100

100

100

100

10 s

100

100

100

100

100

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

0s

100

0

0

0

0

0

0

0

0

0

3s

100

0

0

0

0

3s

0

0

0

0

0

5s

100

90

65

65

65

5s

85

85

85

85

85

7s
10 s

100

100

100

100

100

7s

100

100

100

100

100

100

100

100

100

100

10 s

100

100

100

100

100

48 h

72 h

1 wk

2 wk

A. orisF. nucleatum0s

The percentages of negative achievement rate are indicated（n＝20）.

Table 2
Species and Irradiation

Sterilization test results of paper point（％）

24 h

48 h

72 h

1 wk

2 wk

0

0

0

0

0

0
15

0
0

0
0

0
0

0
0

25

25

25

25

25

5s
7s

10 s

50

50

50

50

50

15 s

60

60

55

55

55

20 s

75

65

65

65

65

30 s

100

100

100

100

100

40 s

100

100

100

100

100

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

0s

0

0

0

0

3s

100

95

65

5s

100

100

7s

100

10 s
Species and Irradiation

B. atrophaeus

0s
3s
5s
7s

S. mutans

A. orisSpecies and Irradiation
E. coli

0

0

0

0

0

100
100

100
100

100
100

100
100

100
100

100

100

100

100

100

10 s

100

100

100

100

100

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

S. intermedius

0s
3s

24 h

0s
3s

0

0

0

0

0

100

100

100

100

100

5s

100

100

100

100

100

7s

100

100

100

100

100

0

10 s

100

100

100

100

100

65

65

100

100

100

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

100

100

100

100

0s

0

0

0

0

0

100

100

100

100

100

3s

100

100

100

100

100

24 h

48 h

72 h

1 wk

2 wk

5s

100

100

100

100

100

7s

100

100

100

100

100

0s

100

0

0

0

0

10 s

100

100

100

100

100

3s

100

35

35

35

35

5s

100

100

100

100

100

7s

100

100

100

100

100

10 s

100

100

100

100

100

F. nucleatum

The percentages of negative achievement rate are indicated（n＝20）.

ODEP Vol. 1, No. 1

18
Sterilization test results of cotton ball（％）

Table 3Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

0s

0

0

0

0

0

3s

0
0

0
0

0
0

0
0

0
0

0

0

0

0

0

10 s

0

0

0

0

15 s
20 s
30 s

0
0

0
0

0
0

0
0

45

45

45

45

45

40 s

100

100

100

100

100

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

0s

0

0

0

0

3s

15

0

0

0

5s

100

45

30

25

25

7s

100

80

65

55

55

10 s

100

100

100

100

100

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

0s

100

0

0

0

0

3s

100

0

0

0

0

5s
7s

100

60

45

45

45

100

95

95

95

95

10 s

100

100

100

100

100

B. atrophaeus

5s
7s

S. mutans

A. orisSpecies and Irradiation
E. coli

24 h

48 h

72 h

1 wk

2 wk

0s

0

0

0

0

0

3s
5s
7s

10
95

10
80

10
80

10
80

10
80

100

100

100

100

100

0

10 s

100

100

100

100

100

0
0

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

S. intermedius

0s
3s

0

0

0

0

0

60

25

25

25

25

5s

95

95

95

95

95

7s

100

100

100

100

100

0

10 s

100

100

100

100

100

0

Species and Irradiation

24 h

48 h

72 h

1 wk

2 wk

0s

0

0

0

0

0

3s

10

0

0

0

0

5s

70

50

50

50

50

7s
10 s

75

70

70

70

70

100

100

100

100

100

F. nucleatum

The percentages of negative achievement rate are indicated（n＝20）.

pressing the irradiation switch.

ting Ⅰ, which is slightly higher than that of conven-

In our prototype sterilizer, the time from pressing

9‒18）
tional sterilizers（200‒280℃）, is that the prototype

the“on”switch to stabilizing the temperature in thesterilizer reflects more of the radiant heat projected

light condensing area was about 20 s. It was reported

onto the sterilized materials compared with the conven-that it takes 9 min for the temperature to stabilize attional type that uses a heating medium, and not all of

230℃ for glass beads, 5 min for salt, and 6 min for mol-the heat energy incident on the materials can be used.

ten metal, which are conventional heating media 13）.

This is also the reason why the core temperature of N.I.

Therefore, the setup time was greatly reduced, and the

dry heat sterilizers using radiation heat has beensterilizer could be used immediately. This feature isincreased to around 360℃ in the past16）.

useful not only in daily clinical practice, but also in

There is no clear ISO standard for simple dry heat

home-visit dental care if the device can be madesterilizers, and each sterilizer is designed with its own

smaller.

optimal temperature. The temperature difference

In addition, it is possible to set the temperature at

between the center of the light condensing area and thethe center of the light condensing area with a control

outer edge of the light condensing area was 80℃.

switch and select setting Ⅰ, which controls the tem-

Although the temperature at the outer edge was notperature by cycling the lamp, or setting Ⅱ, which per-lower than the temperature of a conventional sterilizer,

forms continuously increasing irradiation. The paper

the temperature of the sterilized material may be lower

points and cotton balls, which have poor thermal con-than the apparent temperature for paper and cotton

ductivity, were irradiated with setting Ⅰ, while K-files,

products in a sterilizer with low thermal energy effi-

which are stainless steel and have good thermal con-ciency. Currently, almost no sterilizers available on theductivity, were irradiated with setting Ⅱ.

market can heat K-files up to 330℃, and N.I. sterilizers

The reason why the temperature was 330℃ in set-

have only been reported to heat cotton plugs and paperDec, 2021

Development of a Simple Dry Heat Sterilizer Using Halogen Lamp

points16）.

19

because the number of glove bacteria in the second half

In this study, we observed the surface of the heatedof treatment, just before the working length was deter-

files and performed bending and torsion tests according

mined and provisionally sealed, has been reported to be

to ISO 3630-1 to observe the changes in physical prop-

approximately 1.0‒3.0×103 CFU27）.

erties（Figs. 7 and 8）. Compared with the untreated

For the paper points, negative cultures weregroup, images of the flame-treated group showed metal

obtained after 3‒5 s of irradiation for non-spore-forming

peeling off the surface of the file and the blade disinte-

bacteria, but after 30 s for spore-forming B. atrophaeus.

grating and blunting due to heat, while the halogen

Because the central temperature of the condensing

lamp-irradiated group showed no major changes.

area rises to 330℃, this area seems to be able to kill

It has long been believed that sensitization（a chro-bacteria in a short time. However, the temperaturemium deficiency phenomenon caused by grain precipi-

decreases away from the center, and the temperatures

tation due to chromium carbonization on the surface ofof the ends of the samples（tip of the paper point andthe file）, which is the cause of intergranular corrosion

edge of the cut）located in the outer edge reach onlyin 18-8 stainless steel, occurs when steel is heated for a240‒250℃. Therefore, the spores that survive there

long time at 500‒800℃. In this experiment, the tem-

could germinate.

perature was about 410℃ even after the longest irradi-

Cotton balls were culture negative after 7‒10 s ofation of 15 s, so it is thought that the files were undam-

irradiation for non-spore-forming bacteria, but took 40 s

aged because the temperature was not high enough tofor spore-forming B. atrophaeus. The longer timecause grain precipitation25）.

required for a negative culture compared to the paper

In the bending tests, torque was significantly lower inpoints for all species was thought to be due to the fact

the MANI #15 K-file, and in the torsion tests, torquethat the depth from the surface to the center of the cot-

was significantly lower in the MANI #25 K-file withton ball required more time for heat conduction, allow-

respect to the untreated group. However, in both tests,

ing interior bacteria to survive.

the torque did not decrease significantly compared to

In the K-files, the length of contamination was 16

20）mm, some of which was not placed within the condens-

were met, so the K-files would still be acceptable foring area, and despite the large number of contaminat-

the untreated group, and the ISO standard values
clinical use.

ing bacteria, the culture was negative after 10 s for

There was no significant difference in the fracture

spore-forming B. atrophaeus and after 7 s for other bac-angle among all groups, but one Senseus K-FlexoFile

teria species. This is probably due to the good thermal#40 fractured at an angle under the ISO standard

conductivity of the metal, unlike paper or cotton.

value. The additional 10 pieces tested in accordance

Because it is rare to come into contact with highlywith the standard all exceeded the standard value. The

thermostable bacteria in general dental practice, theresults of these strength tests are in agreement withresults confirmed that clinical sterilization could be

those of Asakura26）, who reported that the mechanicalachieved by irradiating the K-files for 7 s with setting

properties of K-files were not affected until they wereⅡ, and the paper points for 5 s and small cotton balls

heated at 400℃ for 1 h in an electric furnace.

for 10 s with setting Ⅰ at the temperature settings

In the sterilization test, we used B. atrophaeus

used in this study.

（Gram-positive bacteria）, which is an indicator bacte-

The results show that it was able to quickly andrium for dry heat sterilization and has the ability tosafely heat the materials to be sterilized and achieve

form spores; S. mutans, S. intermedius, and A. oris as

sterilization in the light condensing area. This proto-Gram-positive bacteria indigenous to the oral cavity;

type sterilizer had the necessary control boards andand F. nucleatum as Gram-negative bacteria. In addi-

equipment in order to achieve sufficient performance.

tion, E. coli was used as a general Gram-negative bac-

However, since some equipment could be omitted interium. The amount of bacteria applied to the paper

actual clinical applications, it is possible to reduce thepoints and cotton balls was set at 2.0×103 CFU/ml,

size of the sterilizer, and since there is no heating

under the assumption that these materials would be

medium compared to the conventional type, it is possi-

used before provisional sealing at the end of treatment,

ble to reduce the weight. Moreover, the lack of a heat-

ODEP Vol. 1, No. 1

20

ing medium prevented the material from being bent

K-file, there was a tendency that no significant differ-when it was inserted into the sterilization area andence was observed between the conditions with andeliminated contamination of the material by a heating

without irradiation.

medium. The results also suggest that this sterilizer is

5. The time required for 100% of the spore-forming

effective for infected root canal treatment, where abacteria to be sterilized was 10 s for the K-file, 30 s forlarge amount of necrotic pulp or infected dentin is han-the paper point, and 40 s for the cotton ball. For otherdled, or where contamination is suspected due to small

bacterial species, it took 7 s for the K-file, 3 to 5 s for

instruments falling outside the clean area or being

the paper point, and 7 to 10 s for the cotton ball.

exposed to saliva. Moreover, although they do not come

These results suggest that this sterilizer is effective

into direct contact with the sterilized materials, theas used in this experiment, which was conducted underinternal walls of the equipment can be sterilized by

the assumption of clinical procedures.

autoclaving if they are assembled, with the exception of
the halogen lamps, which require wiring, so maintenance is considered possible.

The authors declare no conflicts of interest associatedwith this manuscript.

However, as with conventional sterilizers, it is necessary to insert long instruments, such as tweezers or
K-file handles, that are not heat resistant to direct light
exposure. In addition, sterilization of materials exposed
to a large amount of microorganism contamination will
require a conventional autoclave. The bacterial countset for this study corresponds to that observed during
clinical procedures, and it is therefore desirable to sterilize any suspected contaminated items after cleaning
them with gauze.
In addition, the temperature tended to drop rapidly
away from the center of the light condensing area.
Thus, this instrument is effective for sterilizing small
areas contaminated during use, and conventional methods are required to achieve complete sterilization of theentire instrument. Once used, instruments should only
be used for the same patient, and autoclaves or dryheat sterilizers should be used to sterilize instruments
between patients.

Conclusion
1. In settingⅠin which the central temperature of thecondensing area was maintained, the temperature wasstable at a central of around 330℃ and an outer edge ofaround 250℃ after irradiation for 20 s.
2. In settingⅡwith continuous irradiation, the center
temperature reached 368℃ and the outer edge temperature increased to around 280℃ after irradiation for
10 s.
3. In SEM observation of the irradiated K-file, there
was no change compared to unirradiated file.
4. In the bending test and the torsion test of theReferences
1）Klevens RM, Gorwitz RJ, Collins AS. Methicillin-resistant Staphylococcus aureus: a primer for dentists. J Am
Dent Assoc 2008; 139: 1328‒1337.
2）Volgenant CMC, de Soet JJ. Cross-transmission in thedental office: Does this make you ill? Curr Oral HealthRep 2018; 5: 221‒228.
3）Glick M, Trope M, Bagasra O, Pliskin ME. Human immunodeficiency virus infection of fibroblasts of dental pulp
in seropositive patients. Oral Surg Oral Med Oral
Pathol 1991; 71: 733‒736.
4）Li H, Chen V, Chen Y, Baumgartner JC, Machida CA.
Herpesviruses in endodontic pathoses: association of
Epstein-Barr virus with irreversible pulpitis and apicalperiodontitis. J Endod 2009; 35: 23‒29.
5）Sabeti M, Slots J. Herpesviral-bacterial coinfection inperiapical pathosis. J Endod 2004; 30: 69‒72.
6）Meng L, Hua F, Bian Z. Coronavirus disease 2019
（COVID-19）: Emerging and future challenges for dental
and oral medicine. J Dent Res 2020; 99: 481‒487.
7）Amato A, Caggiano M, Amato M, Moccia G, Capunzo M,
De Caro F. Infection control in dental practice during
the COVID-19 pandemic. Int J Environ Res Public
Health 2020; 17: 4769.
8）Centers for Disease Control and Prevention. Summaryof infection prevention practices in dental settings:
Basic expectations for safe care. Atlanta: Centers for
Disease Control and Prevention, US Dept of Health andHuman Services; October 2016. https://www.cdc.gov/
oralhealth/infectioncontrol/pdf/safe-care2.pdf.（cited
2019. 3. 1）
9）Stewart GG, Williams NB. A preliminary report on theefficacy of molten metal for sterilization of root canal
instruments and materials. Oral Surg Oral Med Oral

Dec, 2021

Development of a Simple Dry Heat Sterilizer Using Halogen Lamp

Pathol 1950; 3: 256‒261.
10）Findlay J. A report on the efficacy of molten metal andball bearings as media for sterilisation. Br Dent J 1955;
98: 318‒323.
11）Grossman LI. Hot salt sterilizer. Br Dent J 1956; 100:283.
12）Oliet S, Sorin S, Brown H. A temperature analysis ofthermostatically controlled root canal sterilizers using
molten metal, glass beads, or salt. Oral Surg Oral Med
Oral Pathol 1958; 11: 37‒42.
13）Yasuoka M, Ideguchi Y, Hiramine K, Osada T. A studyof hot salt sterilizer for root canal instruments. Jpn J
Conserv Dent 1975; 18: 282‒291.（in Japanese）
14）Spring PN. Bacteriologic evaluation of the glass beadsterilizer for endodontics. Oral Surg Oral Med Oral
Pathol 1959; 12: 353‒357.
15）Kitajima K, Eguchi M, Kitano Y, Ezura A, Igarashi M,
Kawasaki K. A study on the efficacy for the sterilization of a newly-designed electric glass bead sterilizer
with digital thermometer. Jpn J Conserv Dent 1993; 36:241‒251.（in Japanese）
16）Tanaka I, Kojima K, Suzuki K, Matsuda J, Nakamura H.
A study of a special electric drying sterilizer. ―An
improved drying apparatus of N.I. system―. Jpn J Conserv Dent 1976; 19: 323‒331.（in Japanese）
17）Tamazawa K, Horiuchi H. A new endodontic sterilizer
using a heated metal slit. Jpn J Conserv Dent 1980; 23:209‒212.（in Japanese）
18）Sawada K, Yazaki M, Kanda M, Suda H, Sunada I. A
study on the efficacy for the sterilization of SL sterilizer. Jpn J Conserv Dent 1981; 24: 246‒251.（in Japanese）
19）International Organization for Standardization. ISO

213630-1: 2019（E）Dentistry―Root-canal instruments―
part 1: General requirements and test methods, 2019.
20）International Organization for Standardization. ISO
3630-5: 2020（E）Dentistry―Endodontic instruments―
part 5: Shaping and cleaning instruments, 2020.

21）International Organization for Standardization. ISO
11138-4: 2017（E）Sterilization of health care products―
Biological indicators―Part 4: Biological indicators fordry heat sterilization processes, 2017.
22）Cisar JO, Vatter AE, Clark WB, Curl SH, Hurst-Calderone S, Sandberg AL. Mutants of Actinomyces viscosus T14V lacking type 1, type 2, or both types of fimbriae. Infect Immun 1988; 56: 2984‒2989.
23）Okahashi N, Sasakawa C, Yoshikawa M, Hamada S,
Koga T. Cloning of a surface protein antigen gene fromserotype c Streptococcus mutans. Mol Microbiol 1989; 3:
221‒228.
24）Bott KF, Davidoff-Abelson R. Altered sporulation andrespiratory patterns in mutants of Bacillus subtilis
induced by acridine orange. J Bacteriol 1966; 92: 229‒
240.
25）Umemura F. Non-destructive evaluation method for
degree of sensitization of stainless steel. J Jpn Weld Soc
1987; 56: 411‒416.
26）Asakura T. The effect of heating temperature for the
cutting efficiency of the K-file for an enlargement of aroot canal. J Fukuoka Dent Coll 2002; 29: 169‒180.
（inJapanese）
27）Niazi SA, Vincer L, Mannocci F. Glove contamination
during endodontic treatment is one of the sources of
nosocomial endodontic Propionibacterium acnes infections. J Endod 2016; 42: 1202‒1211.

ODEP Vol. 1, No. 1

22

Original Article

ODEP 1（1）：22～29，2021

Development of Multilayered Dental Pulp Cell Sheets
and Consideration for Calcification Ability
Katofumi KOYANAGI, Munehiro MAEDA, Miki SEKIYA,
Taro NISHIDA and Masaru IGARASHI
Department of Endodontics, The Nippon Dental University, School of Life Dentistry at Tokyo

Abstract
Purpose: The purpose of this study was to analyze the calcification characteristics of three-dimensional（3D）culture of multi-layered dental pulp cell（DPC）sheets that have potential application in pulp regeneration and evaluation of dental materials and pharmaceuticals.
Methods: DPC were isolated from third molars extracted from three healthy patients aged 18‒30 years
old. DPC sheets were constructed using a temperature-responsive culture dish（UpCell）and a gelatin stamp（Cell Stamp for UpCell）. Histological examination of three-layer and six-layer DPC sheets was performed
using hematoxylin and eosin staining（HE）. DPC sheets on mineral trioxide aggregates（MTA）
were observed
using a scanning electron microscope（SEM）to examine the microstructure. The LOX-1 probe was used to
examine the effects of hypoxia. Calcification ability was analyzed by real-time PCR and Alizarin Red S staining（ARS）. In this study, two-dimensional（2D）culture（monolayer）served as a control.
Results: Thick multilayered cell sheets were confirmed under the macroscopic view and with HE. The
SEM images showed that the DPC sheets covered the MTA. Higher osteo/odontogenic differentiation wasseen in the multi-layered cell sheets than in the monolayer cell sheets. The expression levels of alkaline phosphatase（ALP）, type Ⅰ collagen（COL1A1）, osteocalcin（OCN）, and dentin sialoprotein（DSPP）genes werehigher in the DPC sheets. With ARS, characteristic nodules of mineralization were observed in the cellsheets.
Conclusion: DPC sheets possessed higher mineralizing ability than the monolayer sheets. Cell sheets may
be useful for comparative examination of pharmaceuticals, materials, and pulp regeneration in a 3D environment.
Key words: dental pulp cell（DPC）, osteo/odontogenic differentiation, three-dimensional culture, cell sheets

Corresponding author: Katofumi KOYANAGI, Department of Endodontics, The Nippon Dental University, School of Life Dentistry at Tokyo, 1‒9‒20, Fujimi, Chiyoda-ku, Tokyo 102‒8159, Japan
TEL: ＋81‒3‒3261‒5698, FAX: ＋81‒3‒5216‒3718, E-mail: k-koyanagi_d2117006@tky.ndu.ac.jp
Received for Publication: January 6, 2021/Accepted for Publication: January 15, 2021
DOI: 10.11471/odep.2021‒003

Dec, 2021

Evaluation for Calcification of Cell Sheets

23

at Tokyo with informed consent. DPC was cultured
according to the method described by Huang et al14）.

Introduction

Pulp tissues were minced and digested in a solution
In recent years, three-dimensional（3D）culture has

containing 3 mg/ml collagenase type Ⅰ（Sigma, St.

been performed using various methods. It has been

Louis, MO, USA）and 4 mg/ml dispase（Gibco, Invitro-

reported that 3D culture provides results that closely

gen, Carlsbad, CA, USA）at 37℃ for 60 minutes. Cell

mimic natural tissues as compared to those of two-di-suspensions were obtained using a 70μm strainer andmensional（2D）culture for accurate modeling of living

then seeded in 60 mm culture dishes at 37℃ under 5％organisms1）

and analysis of medicines

2,3）. In dentistry,

3D cultures can be used to assess biocompatibility of

CO2 and cells with passage numbers of 3‒6 were usedin all experiments. The growth medium（GM）used wasand

as follows; minimum essential medium α（MEM-α,periodontal ligament , using scaffolds . However,Gibco）, 10％ fetal bovine serum（FBS, Hyclone, Logan,hypoxia, promotion of calcification in the spheroid struc-

UT, USA）, 100 units/ml penicillin G, 100μg/ml strepto-

dental materials

4,5）

and for regeneration of pulp
7）

6）

8）

9）

ture , and a decrease in viability due to a decrease inmycin（Wako Pure Chemical Industries, Ltd., Osaka,

oxygen and nutrient supply can occur due to the scal-

Japan）, 2 mmol/l L-glutamine（Wako）, and 100μmol/l

10）

ing-up of the 3D structure . Therefore, there may be

L-ascorbicacid-2-phosphate（Sigma）. The osteo/odonto-

underestimation or overestimation of the results whengenic differentiation medium（OM）used was as follows:

evaluating drugs and materials. In view of these facts,

10 nmol/l dexamethasone（Sigma）, 10 mmol/l β-glycer-

we wondered if it would be possible to perform 3D cul-

ophosphate（Sigma）, 50μg/ml L-ascorbic acid phos-

ture that maintains a specified thickness that does notphate（Sigma）, and 10 nmol/l 1,25 dihydroxyvitamin D3

cause hypoxia without using a scaffold and that focuses（Cayman Chemical, Ann Arbor, MI, USA）.

on cell sheet engineering. Cell sheet engineering refers

2 ．Laminating procedure for the DPC sheets

to the use of a temperature-responsive culture dish

DPC sheets were laminated by a modified procedure（UpCell）to collect cell sheets that retain extracellularof the method described by Haraguchi et al15） and thematrix without using enzymes such as trypsin fromnumber of seeded cells was determined with referencemonolayer culture, and to construct 3D structures by

to the manufacturer’s protocol. DPC was seeded at a11）

layering these cell sheets . It is possible to collect cell

cell density of 1.5×105/well into a 24-well UpCell（Cell-

sheets that retain the binding and extracellular matrix

Seed, Tokyo, Japan）and incubated for 5 days in GM,

and stack these cell sheets to construct a 3D struc-then the 7.5％ gelatin stamp（CellStamp for UpCell;

ture11）. In recent years, cell sheet technology has been

CellSeed, Tokyo, Japan）was placed in the well and left

12,13）. However, there are few

at 25℃ for 30 minutes. The cell sheets were trans-studies on the effects of calcification and hypoxia in cellferred to a gelatin gel, which was then transferred tosheets that retain the specified thickness by multi-lay-

the next well. This process was repeated to create mul-ering of dental pulp cell（DPC）sheets using UpCell.

tiple layers. After the stacking was complete, the gela-

Therefore, we used a gelatin stamp（CellStamp fortin gel was heated at 37℃ for 30 min to dissolve theclinically tested in humans

UpCell）to stack cell sheets and analyze the character-gelatin and was washed three times in the medium

istics of DPC sheets for application in pulp regeneration（Fig. 1）.

and evaluation of dental materials and pharmaceuticals.

3 ．Obser vation by hematoxylin and eosin（HE）
staining

Materials and Methods

The monolayer, three-layer, and six-layer cell sheetsin the GM were cultured on a polytetrafluoroethylene

1 ．Cell isolation and culture

membrane for 3 days and fixed with 10％ neutral for-In this study, DPC was collected from three third

malin. Then, the cell sheets were embedded with opti-

molars of three healthy patients（18‒30 years old）who

mal cutting temperature（OCT）frozen compound andhad undergone orthodontic treatment, following a pro-a 7μm section was cut from the OCT blocks andtocol approved by the Institutional Review Board of

stained with HE. HE was observed using an optical

The Nippon Dental University, School of Life Dentistry

microscope.

ODEP Vol. 1, No. 1

24①Preparation of gelatin stamp②For UpCell③Placement on UpCell

・7.5％ gelatin stamp

In case of
6 layers

Repeat3 times④Incubate at 23 ℃ for 30 min⑤24 well plate
Dissolution ofgelatin（37℃，30 min）

⑥In culture
medium3 washes

Used forexperiments

Fig．1 Procedure for laminating cell sheets①Gelatin gel（7.5％）was prepared by molding and placed on a 24‒well UpCell plate. ②‒④Cells were attached to thebottom of the gelatin stamp at 23℃ for 30 min, and the process was repeated three times to make three‒layer sheets.
Six‒layer sheets were prepared by layering a three‒layer sheet on top of another three‒layer sheet after dissolving thegelatin. ⑤Gelatin was dissolved by heating at 37℃ for 30 min in the 24 well plate. ⑥Cell sheets were washed 3 times
with GM.

4 ．Obser vation by scanning electron microscopy（SEM）mean size of calcification（n＝3）.
6 ．Analysis of hypoxia

The monolayer, three-layer, and six-layer cell sheets

Monolayer, three-layer, and six-layer cell sheets werein the GM were cultured on MTA（ProRoot MTA; Den-

cultured in the OM for 3 days, and the hypoxia probe

sply Tulsa Dental Specialties, Johnson City, TN, USA）

solution LOX-1（Medical & Biological Laboratories,

with a diameter of 6 mm for 3 days. The samples were

Aichi, Japan）was used in the medium at a final concen-

fixed with 2.5％ glutaraldehyde solution for 30 min andtration of 2μ/l. The cells were observed under a fluo-

post-fixed with 1％ osmium tetroxide for 2 h at room

rescence microscope（Carl Zeiss LSM 700, Oberkochen,temperature. The samples were then dehydrated withGermany）.

50, 70, 80, 90, 95％, and 99.5％ ethanol for 10 min, followed by replacement with t-butanol and freeze-drying.
The samples were observed using SEM.

7 ．Real-time polymerase chain reaction（PCR）
analysis
Monolayer, three-layer, and six-layer sheets were cul-

5 ．Alizarin Red S staining

tured in the GM and collected at 3, 7, and 14 days.

The monolayer, three-layer, and six-layer cell sheets

Total RNA was extracted using TRIzol reagent

were cultured in the OM for 21 days, washed with

（Thermo Fisher Scientific, Waltham, MA, USA）andphosphate-buffer solution, and fixed with 10％ neutral

Maxwell RSC simplyRNA Cells Kit（Promega Corpora-formaldehyde at room temperature for 15 min. Aftertion, Madison, WI, USA）
according to the manufacturer’s

washing twice with dH2O, 300 ml/well of 40 mmol/l

instructions. Complementary DNA synthesis was per-

ARS（pH 4.1）was added, incubation at room tempera-

formed from total RNA（1μg）using SuperScript VILO

ture was performed for 20 min, then the wells were

Master Mix（Life Technologies, Foster City, CA, USA）.

washed with 1 ml dH2O 4 times for 5 min each. The

Real-time PCR was performed with a THUNDERBIR

samples were observed using an optical microscope andProbe qPCR Mix（TOYOBO, Osaka, Japan）using amacroscopic views. Three random points on the well

StepOnePlus real-time PCR system（Applied Biosys-

were measured by ImageJ software to determine thetems Waltham, MA, USA）under the following condi-Dec, 2021

Evaluation for Calcification of Cell Sheets
Table 1
Genes

25

Sequences of human primers used for real‒time PCR
Upper primers

Lower primers

ALP

5′‒TCCCTGATGTTATGCATGAGC‒3′

5′‒CGAGAGTGAACCATGCCA‒3′

OCN
COL1A1DSPP

5′‒CGCCTGGGTCTCTTCACT‒3′
5′‒TTCTGTACGCAGGTGATTGG‒3′
5′‒TGACACATTTGATCTTGCTAGGA‒3′

5′‒CTCACACTCCTCGCCCTAT‒3′
5′‒GACATGTTCAGCTTTGTGGAC‒3′
5′‒TTTGGGCAGTAGCATGGG‒3′monolayer

three－layer

six－layer

A

B

C

D

E

F

G

H

I

macroscopic
views

Hemotoxylin
and eosin
staining

scanning
electronmicroscope

Fig．2 Histological and SEM findings of cell sheets
Macroscopic views of the monolayer（A）, three‒layer（B）, and six‒layer（C）cell sheets. Hemotoxylin and eosin staining of the monolayer（D）, three‒layer（E）, and six‒layer（F）cell sheets.
Scanning electron microscope view of the monolayer（G）, three‒layer（H）, and six‒layer（I）cell
sheets.
Scale bar＝50 μm

tions: 95℃ for 3 min, followed by 40 cycles at 95℃ for
15 s. The primers for gene amplification were subjected

Results

to 40 cycles at 60℃（45s）for alkaline phosphatase（ALP）, type Ⅰ collagen（COL1A1）and osteocalcin

1 ．Histological analysis（OCN）and at 62℃（45s）for dentin sialoprotein（DSPP）.

The macroscopic view revealed membranous cell

β-actin（NM_001101.2）was used as an endogenous con-sheets with a thickness of three and six layers. The

trol. The primer sequences used are shown in Table 1.

thickness of the monolayer, three-layer, and six-layer

8 ．Statistical Analysis

cell sheets, as seen with HE staining, was approxi-

Statistical significance was analyzed using one-way

mately 7μm, 25μm, and 45μm, respectively. For SEM,

analysis of variance followed by Tukey’s test. Data

when the cell suspension was seeded on the sample, thewere expressed as mean±standard deviation, and p＜

cells were scattered and attached to the MTA, but on

0.05 was considered statistically significant.

the cell sheets, an image covering the sample wasobserved（Fig. 2）.

ODEP Vol. 1, No. 1

26

2 ．Analysis of hypoxia

ization were larger in the six-layer cell sheets（1,573.4±

The monolayer cell sheets showed a few LOX-1 posi-

669.1μm2）than in the three-layer sheets（895.4±269.7

tive cells. More hypoxic cells were observed in the sixlayer cell sheets than in the monolayer and three-layer
cell sheets（Fig. 3）.

μm2）（Fig. 4）.
4 ．Real-time PCR
The expression of ALP was upregulated in the

3 ．Alizarin Red S staining

monolayer, three-layer, and six-layer cell sheets at day 7

Several nodules of mineralization were observed inand downregulated at day 14, but tended to be signifi-

the three-layer and six-layer cell sheets but not in thecantly higher in the three-layer and six-layer cell sheetsmonolayer sheets. In addition, the nodules of mineral-than in the monolayer sheets（p＜0.05）. The expressionof COL1A1 was upregulated in the monolayer, threelayer, and six-layer cell sheets on day 7 and was significantly higher in the six-layer cell sheets than in themonolayer sheets. The expression of OCN was upregulated in the monolayer, three-layer, and six-layer cellsheets at day 14 and was significantly higher in the six-monolayer

three－layer

six－layer

Fig．3 Analysis of hypoxia
24‒well viewed from above hypoxia was detectedwith the LOX‒1 probe in the monolayer, three‒layer,and six‒layer cell sheets.
Scale bar＝100 μm
macroscopic
view

layer（p＜0.05）cell sheets than in the three-layer andmonolayer sheets. The expression of DSPP was upregulated in the monolayer, three-layer, and six-layer cellsheets at day 14 and was significantly higher in the sixlayer（p＜0.05）cell sheets than in the three-layer andmonolayer cell sheets（Fig. 5）.

optical microscope view

A

B

C

D

E

F

G

H

I

monolayer

three－layer

six－layer

Fig．4 Macroscopic and optical views
Macroscopic view of Alizarin Red S staining of the monolayer（A）, three‒layer（D）, and six‒layer（G）cell sheets in the 24‒well plate. Optical microscope view ofthe monolayer（B, C）, three‒layer（E, F）, and six‒layer（H, I）cell sheets. Nodulesof mineralization（indicated with black arrows）can be observed in relation to thethree‒layer and six‒layer cell sheets.
Scale bar＝500μm

Dec, 2021

Evaluation for Calcification of Cell Sheets27

3 days7 days14 days

3 days7 days14 days

3 days7 days14 days

3 days7 days14 days：monolayer：three－layer：six－layer

Fig．5 Real‒time PCR analysis of mineralization related marker genes
Real‒time polymerase chain reaction was performed for ALP, COL1A1, OCN, and DSPP
（n＝3）. The results of two independent experiments performed in triplicate are expressedas the mean（±SD）（＊：p＜0.05）.

Phase contrast imageof DPC
Scale bar=100μm

Fig．S1

Fig．S2 Macroscopic and optical views
Macroscopic view of ARS of the monolayer（A）was cultured in the GM
for 21 days. Optical microscope view of the monolayer（B, C）. Calcification
was not observed（Scale bar=500μm）.

ODEP Vol. 1, No. 1

28

tor-1α（HIF-1α）22）. The effect of hypoxia on cells may

Discussion

cause activation of angiogenesis23）by inducing vascular

In the present study, it was possible to fabricate cellHIF-1α, which may be advantageous for pulp regenera-

sheets to form in membranous by multilayering, astion by transplantation. The above results suggest thatshown by Kikuchi et al16）. In addition, some studies oncell sheet technology may be further developed by

MTA have reported the necessity of direct contact

changing the application depending on the thickness of

between cells and MTA for their differentiation into

the cell sheets.

17）

endothelial growth factor（VEGF）expression related toodontoblast-like cells . Based on these studies, if it isNote that there were some limitations to this study.

possible to cover the MTA as shown by SEM, it may

We were not able to produce stable cell sheets. There-

be possible to analyze the interaction with cells that

fore, it is necessary to consider a more appropriate pro-occurs in the surface layer of the material in more

tocol.

detail and to study the material properties in an envi-

In the future, we plan to conduct a more detailedronment similar to that in vivo. Considering the miner-

study to investigate the material science of multilay-

alization-associated marker gene, after the initialered cell sheets with DPC for regeneration of dental

expression of calcification and in ALP, which is an

pulp.

important factor for mineral deposition18）, downregulation was observed after 14 days in the monolayer

Conclusion

sheets, while the levels were high in the three-layer andsix-layer sheets, which may have been due to the

DPC sheets possessed higher mineralizing ability

occurrence of heterogeneous calcified areas within thethan monolayer sheets. Cell sheets may be useful for

sheets. COL1A1, a major collagen in dentin and bonecomparative examination of pharmaceuticals, materials,

matrix, also showed a similar trend. The expression ofand pulp regeneration in a 3D environment.

OCN, a non-collagenous protein and a marker of osteo/odontogenic differentiation, and DSPP, a representative
marker of odontogenic differentiation, was also higher

The authors have no conflicts of interest directly relevantto the content of this article.

in the multilayered sheets. The expression trends of themarkers were consistent with those seen in previous

Supplemental Date

19）

studies . The above-described upregulation of the mineralization-related marker and increased nodular calcification seen with ARS were observed, especially in the

DPC image（Fig. S1）and ARS staining control（Fig.
S2）were added as Supplemental Date.

six-layer sheets. This may have been due to the thickness of the cells（as seen with HE）, causing the expansion of hypoxic areas and the upregulation of mineralization-associated marker expression20） induced by
localized hyper-calcified areas and hypoxic areas in
ARS. However, when considered in terms of materialogical characterization, since many dental materials
are often compared with cells for their odontogenic
induction21）, a comparative study of calcification-related
factors using six-layer sheets may result in an overestimation of calcification. Therefore, we hypothesized thata 3D culture with about three layers would not beaffected by scaffolding or hypoxia and would allow us toperform comparative material studies in a 3D environment similar to that in vivo. In addition, there is a correlation between LOX-1 and hypoxia-inducible fac-

References
1）Antoni D, Burckel H, Josset E, Noel G. Three-dimensional cell culture: a breakthrough in vivo. Int J Mol Sci
2015; 16: 5517‒5527.
2）Perche F, Torchilin VP. Cancer cell spheroids as a model
to evaluate chemotherapy protocols. Cancer Biol Ther
2012; 13: 1205‒1213.
3）Ravi M, Paramesh V, Kaviya SR, Anuradha E, Solomon
FD. 3D cell culture systems: advantages and applications. J Cell Physiol 2015; 230: 16‒26.
4）Widbiller M, Lindner SR, Buchalla W, Eidt A, Hiller KA,
Schmalz G, Galler KM. Three-dimensional culture ofdental pulp stem cells in direct contact to tricalcium silicate cements. Clin Oral Investig 2016; 20: 237‒246.
5）Pérard M, Le Clerc J, Watrin T, Meary F, Pérez F, Tri-

Dec, 2021

Evaluation for Calcification of Cell Sheets29

cot-Doleux S, Pellen-Mussi P. Spheroid model study

odontoblast-like cells from cultured human dental pulp

comparing the biocompatibility of Biodentine and MTA.
J Mater Sci Mater Med 2013; 24: 1527‒1534.

15）Haraguchi Y, Shimizu T, Sasagawa T, Sekine H, Sakagu-

6）Itoh Y, Sasaki JI, Hashimoto M, Katata C, Hayashi M,
Imazato S. Pulp regeneration by 3-dimensional dental
pulp stem cell constructs. J Dent Res 2018; 97: 1137‒
1143.
7）Raveendran NT, Vaquette C, Meinert C, Ipe DS, Ivanovski S. Optimization of 3D bioprinting of periodontal
ligament cells. Dent Mater 2019; 35: 1683‒1694.
8）Kim NR, Lee DH, Chung PH, Yang HC. Distinct differentiation properties of human dental pulp cells on collagen, gelatin, and chitosan scaffolds. Oral Surg Oral Med
Oral Pathol Oral Radiol Endod 2009; 108: 94‒100.
9）Yamamoto M, Kawashima N, Takashino N, Koizumi Y,
Takimoto K, Suzuki N, Saito M, Suda H. Three-dimensional spheroid culture promotes odonto/osteoblastic
differentiation of dental pulp cells. Arch Oral Biol 2014;
59: 310‒317.
10）Novosel EC, Kleinhans C, Kluger PJ. Vascularization is
the key challenge in tissue engineering. Adv Drug Deliv
Rev 2011; 63: 300‒311.
11）Kobayashi J, Kikuchi A, Aoyagi T, Okano T. Cell sheet
tissue engineering: Cell sheet preparation, harvesting/manipulation, and transplantation. J Biomed Mater Res
A 2019; 107: 955‒967.
12）Nishida K, Yamato M, Hayashida Y, Watanabe K, Yamamoto K, Adachi E, Nagai S, Kikuchi A, Maeda N, Watanabe H, Okano T, Tano Y. Corneal reconstruction withtissue-engineered cell sheets composed of autologous
oral mucosal epithelium. N Engl J Med 2004; 351: 1187‒
1196.
13）Iwata T, Yamato M, Washio K, Yoshida T, Tsumanuma
Y, Yamada A, Onizuka S, Izumi Y, Ando T, Okano T.
Periodontal regeneration with autologous periodontal
ligament-derived cell sheets―a safety and efficacy
study in ten patients. Regen Ther 2018; 9: 38‒44.
14）Huang GT-J, Shagramanova K, Chan SW. Formation ofchi K, Kikuchi T, Sekine W, Sekiya S, Yamato M,
Umezu M, Okano T. Fabrication of functional three-dimensional tissues by stacking cell sheets in vitro. Nat
Protoc 2012; 7: 850‒858.
16）Kikuchi T, Shimizu T, Wada M, Yamato M, Okano T.
Automatic fabrication of 3-dimensional tissues using cellsheet manipulator technique. Biomaterials 2014; 35:2428‒2435.
17）Paranjpe A, Smoot T, Zhang H, Johnson JD. Direct contact with mineral trioxide aggregate activates and differentiates human dental pulp cells. J Endod 2011; 37:1691‒1695.
18）Golub EE, Boesze-Battaglia K. The role of alkaline phosphatase in mineralization. Current opinion in Orthopaedics 2007; 18: 444‒448.
19）Bakopoulou A, Leyhausen G, Volk J, Tsiftsoglou A, Garefis P, Koidis P, Geurtsen W. Comparative analysis of invitro osteo/odontogenic differentiation potential of
human dental pulp stem cells（DPSCs）and stem cells
from the apical papilla（SCAP）. Arch Oral Biol 2011; 56:709‒721.
20）Li L, Zhu YQ, Jiang L, Peng W, Ritchie HH. Hypoxia
promotes mineralization of human dental pulp cells. J
Endod 2011; 37: 799‒802.
21）Emara R, Elhennawy K, Schwendicke F. Effects of calcium silicate cements on dental pulp cells: A systematic
review. J Dent 2018; 77: 18‒36.
22）Zhang S, Hosaka M, Yoshihara T, Negishi K, Iida Y,
Tobita S, Takeuchi T. Phosphorescent light-emitting
iridium complexes serve as a hypoxia-sensing probe fortumor imaging in living animals. Cancer Res 2010; 70:4490‒4498.
23）Amemiya K, Kaneko Y, Muramatsu T, Shimono M,
Inoue T. Pulp cell responses during hypoxia and reoxygenation in vitro. Eur J Oral Sci 2003; 111: 332‒338.

cells on dentin in vitro. J Endod 2006; 32: 1066‒1073.

ODEP Vol. 1, No. 1

30

Original Article

ODEP 1（1）：30～36，2021

Inhibitory Mechanisms of S-PRG Eluate and S-PRG Filler
against Volatilization of Hydrogen Sulfide
Sami OMAGARI1, Nao TANIGUCHI2, 3, Yoshiyuki YOKOGAWA4,
Masahiro YONEDA1, Shigeru YAMAMOTO1, Takashi HANIOKA2
and Takao HIROFUJI1
1）

Department of General Dentistry, Fukuoka Dental College

2）

Department of Preventive and Public Health, Fukuoka Dental College
3）

Oral Medicine Research Center, Fukuoka Dental College

4）

Graduate School of Engineering, Department of Mechanical and Physical Engineering, Osaka City University

Abstract
Purpose: The aim of this study was to elucidate the inhibitory mechanism of a surface pre-reacted
glass-ionomer（S-PRG）eluate and S-PRG filler against volatilization of hydrogen sulfide（H2S）.
Methods: The immediate inhibitory activity of S-PRG eluate on H2S volatilization was assessed by mixing
with a solution of sodium hydrogen sulfide（NaHS）in equal amounts for 5 min, and measuring the gaseousconcentration of H2S by gas chromatography. Subsequently, changes in the concentration of H2S in the gaseous and liquid phases, as well changes in the pH in the liquid phase, were examined following the addition of
S-PRG eluate, S-PRG filler, spherical silica, and potassium bicarbonate（KHCO3）.
Results: S-PRG eluate inhibited 31.6％ of H2S volatilization in the reaction mixed with NaHS solution but
with no significant difference（p＝0.065）. Following the addition of S-PRG eluate to the H2S solution, the concentration of H2S was reduced by 20％ in the gaseous phase and increased by 5％ in the liquid phase. There
was no change in the gaseous concentration of H2S in the reaction supplemented with spherical silica,
whereas the gaseous concentration of H2S was reduced in the reaction supplemented with KHCO3, similar tothat with S-PRG filler. The pH of the liquid phase was 5.77 in the H2S solution, 6.01 in that with spherical silica, 7.07 in that with S-PRG filler, and 8.50 in that with KHCO3.
Conclusion: The results of this study suggest that the inhibitory effect of S-PRG eluate and S-PRG filler on
H2S volatilization might be due to their buffering ability.
Key words: buffering ability, hydrogen sulfide, oral malodor, S-PRG filler

Corresponding author: Nao TANIGUCHI, Department of Preventive and Public Health, Fukuoka Dental College, 2‒15‒1,
Tamura, Sawara-ku, Fukuoka 814‒0193, Japan
TEL: ＋81‒92‒801‒0411, FAX: ＋81‒92‒801‒4909, E-mail: naojsz@college.fdcnet.ac.jp
Received for Publication: January 15, 2021/Accepted for Publication: March 3, 2021
DOI: 10.11471/odep.2021‒004

Dec, 2021

Inhibition Effect of S-PRG on Oral Malodor

31

givalis and Fusobacterium nucleatum14）, to inhibit biofilm formation and disrupt mature biofilms; however, its

Introduction

anti-bacterial activity was limited15）. In further clinical
Oral malodor, or bad breath, is one of the major com-

trials, oral rinsing and tongue cleaning with S-PRG elu-

plaints from patients visiting dental clinics, behind only

ate reduced oral malodor compared to the control15）.

dental caries and periodontitis. Clinical causes of oral

Since the reduction of oral malodor occurred immedi-

malodor include periodontitis, poor oral hygiene, tongue

ately after oral rinsing with S-PRG eluate, it is thought

debris, deep caries, inadequately fitted restorations,that S-PRG eluate suppresses oral malodor by an imme-

1‒4）

endodontic lesions, and low salivary flow

. The major

diate mechanism that does not involve anti-biofilm,

compounds involved in oral malodor are volatile sulfur

anti-bacterial, or anti-enzyme actions. The purpose ofcompounds（VSCs）, and approximately 90％ of thethis study was to investigate whether S-PRG eluate andVSCs in mouth air are hydrogen sulfide（H2 S）and

S-PRG filler suppress oral malodor independent of their

methyl mercaptan（CH3SH）5）. VSCs are produced byeffect on oral bacteria, and to clarify the underlying

oral bacteria in the oral cavity during metabolism of themechanism.

sulfur-containing amino acids cysteine and methionine6）.
Gram-negative anaerobes are important producers ofMaterials and Methods

VSCs7‒9）. Persson et al.7） evaluated the capacity of 163
culturable oral bacterial strains to produce VSCs; all

1 ．Preparation of S-PRG eluate and S-PRG filler

bacterial strains tested formed H2S from L-cysteine,

The S-PRG eluate（lot 031501K; Shofu Inc., Kyoto,whereas less than one-third of the strains produced

Japan）and S-PRG filler（lot 120501; Shofu Inc.）used in

CH3SH from L-methionine. Periodontal pathogens iso-

this study were generated as previously described18,19）.

lated from subgingival plaque, such as Porphyromonas

S-PRG eluate was prepared with S-PRG filler under thegingivalis, Prevotella intermedia, Tannerella forsythia,

specific conditions that produce the highest concentra-

and Treponema denticola, have been shown to generatetion of each ion19）. Elemental analysis of ions released

significant amounts of H2S and CH3SH8）. An examina-

from S-PRG filler（Al, B, Na, Si, and Sr）was performed

tion of the microbiota composition in tongue biofilms ofusing inductively coupled plasma atomic emission spec-

individuals without periodontitis, or only a slight degree

troscopy（ICP-AES; ICPS-8000, Shimadzu Co., Kyoto,of periodontitis, suggested that the major species ofJapan）and fluoride ion electrodes（fluoride electrode,

H2S-producing bacteria are Veillonella, Actinomyces,

Model 9609BN; pH/ion meter, Model 720A; Orionand Prevotella9）.

14）
Research Inc., MA, USA）
. The ion concentrations of

Dental restorative material containing surface pre-re-

S-PRG eluate used to evaluate the immediate inhibitory

acted glass-ionomer（S-PRG）filler has recently beeneffect of S-PRG eluate on gaseous H2S were as follows:shown to possess both anti-dental plaque and anti-bac-

Al, 1.5 ppm; B, 890.5 ppm; Na, 368.2 ppm; Si, 6.0 ppm; Sr,terial properties10‒15）. S-PRG filler can both release and78.4 ppm; and F－, 105.0 ppm.

recharge fluoride ions（F）, as well as release other ions,
including strontium（Sr）, silicon（Si）, boron（B）, sodium

2 ．Immediate inhibitor y ef fect of S-PRG eluate
on gaseous H2S

16）（Na）, and aluminum（Al）. S-PRG filler and released

Gaseous H2 S was prepared from a dilute aqueous

ions have anti-demineralization and re-mineralization

solution of sodium hydrogen sulfide（NaHS）. Chlorides

effects17,18）. Previous studies concerning the anti-cario-of two metal ions released from S-PRG filler, AlCl3 andgenicity of S-PRG filler have reported that the eluate of

SrCl 2 , were prepared as 1 mol/l aqueous solutions.

S-PRG filler inhibits bacterial growth, adhesion, andThen, 2 ml of S-PRG eluate, 1 mol/l AlCl3 solution, or 1

gene expression related to sugar metabolism of Strepto10,12）

coccus mutans

mol/l SrCl2 solution was added to 2 ml of 10－5％ NaHS

. We have been studying the effect ofsolution in 15 ml tubes, which were sealed and incu-

S-PRG eluate on oral microbiota including periodonto-bated at room temperature for 5 min. Then, 1 ml of thepathic bacteria14,15）. S-PRG eluate has been shown togas phase was collected and analyzed using a gas chro-suppress the protease and gelatinase activities of P.

matograph（model GC2014; Shimadzu Co.）
. Distilled

gingivalis, as well as the coaggregation between P. gin-water（d-water）and 1 mol/l ZnCl2 solution were usedODEP Vol. 1, No. 1

32

H2S

Dilute sulfuric acid

Add S－PRG filler or S－PRG elute,spherical silica, KHCO3

Gaseous phases

Neutralizer（NaOHaq）

H2S stock solution

Ferric sulfide

Measured by gas
chromatography
Liquid phases

A
Fig. 1

B

Hydrogen sulfide（H2S）gas generator（A）and equipment for preparation of the H2S solution（B）

as negative and positive controls, respectively. All test

5 ．Changes in H2S concentration in the gaseous

reagents were obtained from FUJIFILM Wako Purephase and pH in the liquid phase following

Chemical Co.（Osaka, Japan）. The experiments wereaddition of S-PRG filler

repeated at least three times.

The H2S concentration in the gaseous phase and pH

3 ．Preparation of H2S solution

in the liquid phase were measured at 0, 0.5, 1, 2, and 3 h

H2S gas was produced by adding dilute sulfuric acidfollowing the addition of 0.1 g S-PRG filler, 0.1 g spheri-through a dropping funnel into a four-necked flask con-cal silica, or 0.1 g potassium bicarbonate（KHCO3）to 100

taining ferric sulfide. H2S gas was transferred into an

ml of H2S solution（500 ppb）in an Erlenmeyer flask.

Erlenmeyer flask and bubbled through 150 ml of Milli-Q

6 ．Changes in H2 S concentration in the liquidwater to obtain a stock solution of H2S（Figure 1A）.

phase following addition of S-PRG filler by the

The H2S stock solution was removed by syringe andmethylene blue method

transferred to an Erlenmeyer flask containing 100 ml

In a previous study, S-PRG filler was added to H2SMilli-Q water, and rotated at 400 rpm for 1 h to obtain asolution and the changes in H2S concentration in thesolution of H2S with a concentration of 500 ppb（Figure

liquid phase were examined by the methylene blue

1B）. Finally, 2μl of the liquid and 30μl of the gas in themethod20）. Immediately following the addition of S-PRG

headspace of the H2S solution were tested by gas chro-

filler in the above experiment（initial）, as well as at 0.5,

matography to confirm that the concentration of H2S

1, 2, and 3 h, a 0.1 ml sample was taken from the liquid

was 500 ppb.

phase as a test solution. Briefly, 0.1 ml test solution was

4 ．Changes in H2S concentration in the gaseous

added to 10 ml distilled water in a 15 ml stoppered test

and liquid phases following addition of S-PRG

tube; 1 ml p-aminodimethylaniline dichloride（10 mmol/l

eluatein 10 N H2SO4）
and 1 ml FeCl（20mmol/l in 1％ H2SO4）
3

The changes in H2S concentration in the gaseous andwere also added（Figure 2）. After incubation at room

liquid phases following addition of 0.5 ml S-PRG eluate

temperature for 15 min, the absorbance was measured

to 100 ml of H2S solution（500 ppb）in an Erlenmeyer

at 665 nm using an ultraviolet-visible-near-infrared

flask were measured by gas chromatography（GC-14B;

spectrophotometer（V-550; Jasco Co., Tokyo, Japan）.

Shimadzu Co.）at 0, 1, 2, 3, and 4 h. Then, 30μl was

Measurement of only the reagent without the test solu-

sampled from the headspace as a gaseous phase sam-tion was used as the baseline.

ple, 2μl was sampled from the reaction mixture as a

7 ．Statistical analysis

liquid phase sample, and the H2 S concentration was

Welch’s t-test was used to evaluate the immediate

measured.

inhibitory effects of metal ions and S-PRG eluate on H2S
volatilization compared to the addition of d-water（negative control）. Differences were considered to be significant at p＜0.05. Statistical analyses were carried out

Dec, 2021

Inhibition Effect of S-PRG on Oral Malodor
NH2

H3C

Fe3＋, H＋Oxidation
H3C

HCl

N

Cl－

CH3

N
CH3

N＋

N

CH3

CH3

H2S
Reduction＋NH4Cl＋H＋

33
N
H3C

S

N
CH3

Methylene blue

Dimethyl－p－phenylenediamine

Cl－
N＋

CH3
＋4H＋

CH3

λmax＝665nm

Fig. 2 Methylene blue method
Under acidic conditions, H2S produces methylene blue by the reaction of p-aminodimethylaniline hydrochloride

with iron（Ⅲ）chloride as an oxidizing agent. Since methylene blue has a maximum absorption wavelength of 665

10
9
8
7
6
5
4
3
2
1
0

Control S－PRG
eluateFig. 3

SrCl2

AlCl3

＊
ZnCl2

Immediate inhibitory effect of surface
pre-reacted glass-ionomer（S-PRG）eluate on gaseous H2S

Bars show mean±standard error. S-PRG eluate inhibited 31.6％ of H2S volatilization（p＝0.065）, while AlCl3 and SrCl2 did not inhibit H2S
volatilization. The aqueous solution of ZnCl2

almost completely inhibited H2S volatilization（＊：p＜0.001）.

Relative concentrations of H2S
in both phase

Concentration of H2S（ppm）

nm, it is possible to quantify H2S using absorptiometry.

1.2
1.0
0.8
0.6
Gaseous phase

0.4

Liquid phase

0.2
0

0

1

2

3

Reaction time（hour）after adding S－PRG Eluate
Changes in H2S concentration in the gaseous andliquid phases following addition of S-PRG eluate（representative data）
The concentration of H2S in the gaseous phase（filled

Fig. 4circles）decreased by 20％, while that in the liquid phase（filled squares）increased by 5％.

2 ．Changes in H2S concentration in the gaseousand liquid phases, and pH, following additionof S-PRG eluate or S-PRG filler

using the R software package（ver. 3.6.1.; http://www.

Although the reproducibility was confirmed byrepeated experiments, the reaction was easily affected

R-project.org）.

by the temperature and the measured values varied,

Results

and thus representative data are shown. One hour after
S-PRG eluate was added to the H2S solution（500 ppb）

1 ．Direct inhibitor y ef fect of S-PRG eluate on
gaseous H2S

and stirred at 400 rpm, the concentration of H2S in thegaseous phase decreased by 20％, while that in the liq-

In the reaction mixed with the 10－5％ NaHS solution,

uid phase increased by 5％（Figure 4）; these concentra-

neither aqueous solutions of AlCl3（－9.68％）nor SrCl2

tions remained unchanged after 4 h. Following the addi-（－4.63％）inhibited H2S volatilization, whereas S-PRG

tion of S-PRG filler, the concentration of H2S in the gas-

eluate inhibited H2S volatilization by 31.6％（p＝0.065）

eous phase decreased by approximately 70％ after 1 h,

（Figure 3）. The aqueous solution of ZnCl2 almost com-and by approximately 80％ after 3 h（Figure 5）. As

pletely inhibited H2S volatilization（99.9％, p＜0.001）.

shown in Figure 5, the reaction with KHCO3 resulted ina similar decrease in the concentration of H2S in thegaseous phase as that following the addition of S-PRG
filler. By contrast, the addition of spherical silica did not

ODEP Vol. 1, No. 1

1.2
1.0

pH＝6.01

pH＝5.77

0.8

S－PRG filler
Spherical silica

0.6

KHCO3

0.4

pH＝7.070.2
0Abundance ration in solution

Relative concentration of H2S
in gaseous phase34

pH＝8.5001

2

3

Reaction time（hour）after adding each filler
Changes in H2S concentration in the gaseous andliquid phases, and in pH, following addition of
S-PRG filler（filled circles）
The concentration of H 2 S in the gaseous phaseFig. 5

1.0
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0

H＋＋SH－（free）
H2S
pKa1＝6.88

0

2

4

6

H2S
（free）
Fig. 7decreased by 80％ after 3 h. The reaction with KHCO3（filled triangles）
resulted in a decrease in H2S concentration in the gaseous phase similar to that following addition of S-PRG filler. The addition of spherical silica（filledsquares）did not affect the concentration of H2S in the

pKa2≒14.15

8pH
H＋＋SH－

10

12

14

2H＋＋S2－

Relative abundance of each dissociated speciesof H2S at each pH

gaseous phase. Representative data are shown.

A

0.020

Abs.

0.015
0.010
0.005
0.000

400

450

500

550

600

650

Wavelength（nm）
B

Relative concentration of H2S

initial

0.5h

1h

2h

700
665 nm

3h

750

800

Baseline

1.4
1.2
1.0
0.8
0.6
0.4
0.2
0
0.0

Reaction Timeinitial

0.5h

1h

2h

3h

Relative concentration of H2S

1.000

1.159

1.253

1.194

1.258

1.5

2.0

2.5

0.5

1.0

3.0

3.5

Reaction time（hour）
Fig. 6 Effect of addition of S-PRG filler on H2S concentration in liquid phase（A）Changes in H2S concentration in the liquid phase following addition of

S-PRG filler by the methylene blue method.（B）Changes in relative concentration of H2S in the liquid phase measured by the absorbance at 665 nm.
Representative data are shown.

affect the concentration of H2S in the gaseous phase.

spherical silica, were 5.77, 7.07, 8.50, and 6.01, respec-

The pH values in the liquid phase prior to the reaction,

tively. The concentration of H2S in the liquid phase wasand 3 h after the addition of S-PRG filler, KHCO3, andexamined with the methylene blue method（Figure 6

Dec, 2021

Inhibition Effect of S-PRG on Oral Malodor

35

A）. The relative concentration of H2 S in the liquid

solution rises, and H2S（liquid phase: free）in the liquid

phase increased by 15.9％, 25.3％, 19.4％, and 25.8％ atphase may decrease. Since the H2S aqueous solution is0.5, 1, 2, and 3 h after the addition of S-PRG filler（Fig-weakly acidic, H2S（liquid phase: free）＞H＋＋SH－ in theure 6B）.

liquid phase before the material is charged, but when
the pH of the liquid phase is＞6.88, H2S（liquid phase:free）＜H＋ ＋SH－. In other words, H2 S（liquid phase:

Discussionfree）in the liquid phase decreases in an environment
We previously reported that metal ions such as Ag＋,
2＋

2＋

2＋

Cd , Cu , and Zn

inhibit H2S volatilization by chem-

ical binding21）. In both previous and current experi2＋

3＋

with pH＞6.88, and H2S in the gas phase may move tothe liquid phase. The concentration of H2S in the gas
and liquid phases was examined every hour for 3‒4 h,

did not inhibit volatilization ofand the reaction was nearly complete after 1 h follow-

H2S. On the other hand, S-PRG eluate inhibited 31.6％ of

ing the addition of both S-PRG filler and S-PRG eluate.

ments, Srand AlH2 S volatilization in the reaction mixed with 10－5％

The reaction occurred rapidly, as the concentration of

NaHS. S-PRG eluate immediately inhibited H2S volatil-

H2S in the gas phase decreased by 31.6％ after 5 min in

ization by a mechanism distinct from its action on bac-the mixing experiment with NaHS. The concentration

teria and chemical binding by metal ions. In terms ofof H2S in the gas phase is believed to be lower in S-PRG

the anti-bacterial action of S-PRG filler on S. mutans, itfiller than in S-PRG eluate, because the S-PRG filler con-has been reported that borate and fluoride ions aretinued to release ions to maintain the buffer capacity.

among the ions released from the filler22）. Sr2＋ are also
often taken up by enamel and are considered to beConclusion

involved in the inhibitory effect of S-PRG eluate on thedemineralization of enamel23）. In that study, the absorp-

In conclusion, the weak inhibitory effect of S-PRG

tion of multiple ions from S-PRG filler eluate into theeluate and S-PRG filler on H2S volatilization might be

tooth structure occurred much faster than that of eachdue to their buffering ability. Oral malodor might be

ion alone. As described above, S-PRG filler is character-

suppressed by complex mechanisms of S-PRG eluate

ized by its ability to allow the released ions to act inand filler that include anti-biofilm, anti-bacterial, andconcert with multiple ions, rather than acting alone.

anti-enzyme actions against bacteria as well as a

However, we conclude that suppression of the volatil-

non-bacterial buffer action.

ization of H2S does not occur via a chemical bond, sinceno reaction occurs with the ions alone.
The pH of S-PRG eluate is approximately 7.8

Ethical statement19）.

Not applicable.

Therefore, we focused on the buffering effect of S-PRG
filler and S-PRG eluate. S-PRG filler, KHCO3, and spheri-Acknowledgments

cal silica were added to an aqueous solution of H2S, andWe would like to thank Mr. Toshiyuki Nakatsuka and Mr.
Daisuke Hara of Shofu Inc.
（Kyoto, Japan）for their help anddiscussions. We would also like to thank Textcheck（www.
textcheck.com）for English language editing. This study was
supported in part by a Grant-in-Aid for Scientific Research（No. 26463175）from the Ministry of Education, Culture,
Sports, Science and Technology（MEXT）, Japan, and theOral Medicine Research Center of Fukuoka Dental College.

the concentration of H2S in the gas phase and pH in theliquid phase were measured. The concentration of H2S
in the gas and liquid phases respectively decreased andincreased when KHCO3 and S-PRG filler were added,
which makes the aqueous solution weakly alkaline. The
effects of pH were considered in these experiments. In
the reaction vessel, the gas-liquid and acid dissociation
equilibria are in equilibrium（Figure 7）:
H2S（gas phase）⇄H2S（liquid phase: free）⇄H＋＋SH－
Also, when limited to solution only,
H2S（liquid phase）⇄H＋＋SH－

Since its acid dissociation constant（pKa1）is 6.88, theequilibrium tends to dissociate ions when the pH of the

Conflict of interest
The authors declare that they have no competing interest.

ODEP Vol. 1, No. 1

36
cus mutans. Sci Rep 2018; 8: 5056.

References
1）Suzuki N, Fujimoto A, Yoneda M, Watanabe T, Hirofuji
T, Hanioka T. Resting salivary flow independently associated with oral malodor. BMC Oral Health 2016; 17: 23.
2）Morita M, Wang HL. Association between oral malodor
and adult periodontitis: a review. J Clin Periodontol
2001; 28: 13‒19.
3）Yoneda M, Naito T, Suzuki N, Yoshikane T, Hirofuji T.
Oral malodor associated with internal resorption. J OralSci 2006; 48: 89‒92.
4）Garret NR. Poor oral hygiene, wearing dentures at night,
perceptions of mouth dryness and burning, and lower
educational level may be related to oral malodor in denture wearers. J Evid Based Dent Pract 2010; 10: 67‒69.
5）Tonzetich J. Direct gas chromatographic analysis of sulphur compounds in mouth air in man. Arch Oral Biol
1971; 16: 587‒597.
6）Scully C, Porter S, Greenman J. What to do about halitosis. BMJ 1994; 308: 217‒218.
7）Persson S, Edlund MB, Claesson R, Carlsson J. The formation of hydrogen sulfide and methyl mercaptan byoral bacteria. Oral Microbiol Immnuol 1990; 5: 195‒201.
8）Suzuki N, Yoneda M, Takeshita T, Hirofuji T, Hanioka T.
Induction and inhibition of oral malodor. Mol Oral
Microbiol 2019; 34: 85‒96.
9）Washio J, Sato T, Koseki T, Takahashi N. Hydrogen sulfide-producing bacteria in tongue biofilm and their relationship with oral malodor. J Med Microbiol 2005; 54:889‒895.
10）Saku S, Kotabe H, Scougall-Vilchis RJ, Ohashi S, Hotta M,
Horiuchi S, Hamada K, Asaoka K, Tanaka E, Yamamoto
K. Antibacterial activity of composite resin withglass-ionomer filler particles. Dent Mater J 2010; 29:
193‒198.
11）Han K, Takenaka S, Okuji T. Evaluation of selected
properties of a prototype S-PRG filler containing root
canal sealer. J Conserv Dent 2007; 50: 713‒720.（in Japanese）
12）Nomura R, Morita Y, Matayoshi S, Nakano K. Inhibitory
effect of surface pre-reacted glass-ionomer（S-PRG）
eluate against adhesion and colonization by Streptococ-

13）Takakusaki K, Fueki K, Tsutsumi C, Tsutsumi Y, Iwasaki N, Hanawa T, Takahashi H, Takakuda K, Wakabayashi N. Effect of incorporation of surface pre-reacted
glass ionomer filler in tissue conditioner on the inhibition of Candida albicans adhesion. Dent Mater J 2018; 8:453‒459.
14）Yoneda M, Suzuki N, Masuo Y, Fujimoto A, Iha K,
Yamada K, Iwamoto T, Hirofuji T. Effect of S-PRG eluate on biofilm formation and enzyme activity of oral
bacteria. Int J Dent 2012; 2012: 814913.
15）Suzuki N, Yoneda M, Haruna K, Masuo Y, Nishihara T,
Nakanishi K, Yamada K, Fujimoto A, Hirofuji T. Effectsof S-PRG eluate on oral biofilm and oral malodor. Arch
Oral Biol 2014; 59: 407‒413.
16）Nakatuka T, Yasuda Y, Kimoto K, Mizuno M, Negoro N.
Dental fillers. United States Patent No. 6620861; 2003.
17）Shiiya T, Mukai Y, Tomiyama K, Teranaka T. Anti-demineralization effect of a novel fluoride-releasing varnish on dentin. Am J Dent 2012; 25: 347‒350.
18）Ito S, Iijima M, Hashimoto M, Tsukamoto N, Mizoguchi
I, Saito T. Effects of surface pre-reacted glass-ionomer
fillers on mineral induction by phosphoprotein. J Dent
2011; 39: 72‒79.
19）Fujimoto Y, Iwasa M, Murayama R, Miyazaki M, Nagafuji A, Nakatsuka T. Detection of ions released from
S-PRG fillers and their modulation effect. Dent Mater J
2010; 29: 392‒397.
20）Yano M. Quantification of hydrogen sulfide in hot springwater using methylene blue absorptiometry. Annual
Report of the Hyogo Prefectural Institute of Public
Health and Environmental Sciences 2002; 1: 181‒186.
21）Suzuki N, Nakano Y, Watanabe T, Yoneda M, Hirofuji T,
Hanioka T. Two mechanisms of oral malodor inhibition
by zinc ions. J Appl Oral Sci 2018; 26: e20170161.
22）Miki S, Kitagawa H, Kitagawa R, Kiba W, Hayashi M,
Imazato S. Antibacterial activity of resin composites
containing surface pre-reacted glass-ionomer（S-PRG）
filler. Dent Mater 2016; 32: 1095‒1102.
23）Ogawa A, Wada T, Mori Y, Uo M. Time dependence
multi-ion absorption into human enamel from surface
pre-reacted glass-ionomer（S-PRG）filler eluate. Dent
Mater J 2019; 38: 707‒712.

Dec, 2021

37

Original Article

ODEP 1（1）：37～43，2021

Clinical Assessment of Resin-coating Technique for Dentin
after Cavity Preparation
Hanemi TSURUTA, Shusuke KUSAKABE,
Michael F BURROW＊ and Toru NIKAIDO
Department of Operative Dentistry, Division of Oral Functional Science and Rehabilitation,
Asahi University School of Dentistry＊

Restorative Dental Sciences, Faculty of Dentistry, University of Hong Kong

Abstract
Purpose: Many studies have been conducted on the resin-coating technique for indirect restorations, andits efficacy has been reported. This study investigated the clinical course of cases with the resin-coating
technique for dentin after inlay cavity preparation and examined the efficacy of the technique clinically.
Methods: Twenty cases in 18 patients visiting one of three dental clinics that were collaborating with thisinvestigation between September 2014 and October 2019 were evaluated. The patients were treated using
the resin-coating technique by applying a thin coating material（Hybrid Coat Ⅱ, HC Ⅱ）after inlay cavity
preparation. Patient dental and medical records as well as clinical information that included tooth site, procedures, materials used, and clinical symptoms before, during, and after the procedures and at the time ofrecall were obtained using set clinical evaluation criteria.
Results: Fifty percent of the restorations were performed in premolars and 50％ in molars. Local anesthesia was used in all cases except one. Resin coating was performed using HC Ⅱ alone in 40％ of cases and HC
Ⅱ combined with a flowable resin in 60％ of cases. After inlay placement, 95％ of the cases（19 cases）had afavorable outcome. Of the 10 cases（50％）where the patient experienced pain on cold water stimulation
before treatment, nine cases had a favorable outcome; however, in one case pain on drinking cold water persisted and the patient then presented with pulpal symptoms, thereby requiring a pulpectomy.
Conclusion: The results demonstrate the clinical efficacy of the resin-coating technique using HC Ⅱ on
inlay cavity preparations.
Key words: resin-coating technique, Hybrid Coat Ⅱ, inlay cavity preparation

Corresponding author: Shusuke KUSAKABE, Department of Operative Dentistry, Division of Oral Functional Science andRehabilitation, Asahi University School of Dentistry, 1851, Hozumi, Mizuho, Gifu 501‒0296, Japan
TEL & FAX: ＋81‒58‒329‒1442, E-mail: kusakabe@dent.asahi-u.ac.jp
Received for Publication: May 15, 2021/Accepted for Publication: August 11, 2021
DOI: 10.11471/odep.2021‒005

ODEP Vol. 1, No. 1

38

resin coating is a reliable treatment21）. However, noclinical study has evaluated the resin coating for cavity

Introduction

preparations. Therefore, this retrospective study invesIn recent years, the use of direct composite resto-

tigated the clinical performance of inlay restorations

rations in molars has improved with the advancement

placed using a resin-coating technique in collaborationof adhesive materials and technologies. Currently, awith three dentists who routinely employ the res-composite restoration can be promptly performed byin-coating technique for vital teeth, and assessed the

preparing the cavity such that only the‘caries-infected’validity and efficacy of the procedure.

dentin is removed and‘caries-affected’dentin is preserved, thereby protecting the remaining dentin andMaterials and Methods

pulp. However, for extensive carious lesions in molars
where the restoration is subjected to greater occlusal

1 ．Materials for resin coating

loading and the restorations may need to protect cusps,

The composition of Hybrid Coat Ⅱ（HC Ⅱ Sun Med-an indirect restoration such as an inlay or onlay can beical, Moriyama, Japan）, a resin coating material used inselected. Generally, the degree of dentin exposure after

this study, is shown in Table 1. HC Ⅱ is the next ver-preparation for an indirect restoration is much greater.

sion of HC（Sun Medical）, which has been available in

The resin-coating technique was introduced based on

the market since January 2010. HC Ⅱ was developed

the idea of direct composite resin restorations and theon the concept of a one-step bonding agent, which con-application of a reliable adhesive system for indirect

sists of a one-bottle bonding agent and a dedicatedrestorations. The resin-coating technique is used tosponge22）. When HC Ⅱ is used, a dedicated sponge con-cover the dentinal surface exposed after cavity prepa-

taining adhesion promoters is used to apply the bonding

ration for an indirect restoration such as an inlay ormaterial to the tooth surface. In Japan, HC Ⅱ has beencrown with an adhesive system for protecting thecovered under the Japanese national health insurance

exposed dentin and pulp1‒4）. Specifically, coating thesystem since December 2019 as a material for resin

exposed dentinal surface immediately before impres-coating of prepared surfaces of vital abutment teeth.

sion-taking with an adhesive system aims to form a thin

2 ．Methods for investigation

film on the dentinal surface which helps protect theThis study was approved by the Ethics Committee ofpulp and improves adhesion of the resin cement to the

Asahi University School of Dentistry（approval number

dentin, thereby improving the marginal seal1）. By using

32007）. The study was conducted in collaboration withthe resin-coating technique, few adhesive failures of res-three dentists in private practice who frequently per-torations at the dentinal interface occur due to theform vital tooth inlay restorations using the resin-coat-

improvement in the adhesion of the resin cement to theing technique. The outline of the clinical study and its

dentin. This also leads to a decrease in the frequency of

methods were explained to the dentists and their con-marginal and bacterial leakage which has been

sent to join the study was obtained.

2 , 5‒1 3）. Conservation of tooth structure,

The study included 18 patients（ten males, eightdecreased patient discomfort, and a favorable clinicalfemales; mean age, 51.4 years old）visiting one of thecourse of indirect restorations have also been

three dentists between September 2014 and October

reportedreported

1,2,14‒19）

2019. Patient records were identified for those who.

In Japan, the resin-coating technique for preparing

received an inlay for a vital tooth. The age of thevital abutment teeth has been covered under the Japa-

patients is shown in Table 2. Nineteen cases in 18

nese national health insurance system since December

patients who could be followed up after the restoration

2019 and its use is expected to become more common.

and who also consented to join the investigation wereOn the other hand, the clinical application of the res-investigated.

20）

in-coating technique is still being improved . Labora-

Immediately after the cavity preparation, the pre-tory studies have demonstrated its superior perfor-

pared tooth was resin-coated using HC Ⅱ followed bymance; a clinical study on the resin-coating techniqueimpression-taking. At the next visit, an inlay was placedfor crown preparation of vital teeth reported that thewith a resin cement.

Dec, 2021

Clinical Assessment of Resin-coating Technique
Table 1

Brand
Hybrid Coat Ⅱ（HC Ⅱ）

39

Composition and usage of resin-coating material

Manufacturer

Composition

Sun Medical（Moriyama, Japan）

Procedure

liquid：acetone, MMA, 4-META others, water ①Liquid stirring and mixing with acoat sponge/micro-brush sponge
Powder in coat sponge/micro-brush sponge： ②After applying the mixed solution
aromatic amine, aromatic sulfinate
for 10 s, air-blowing for 5 s③Light irradiation for 5 s

MMA：methyl methacrylate, 4-META：4-methacryloxyethyl trimellitate anhydride

Table 2Sex

Category
Number of cases（Percentage）

Baseline information of the participants

Male

Age（yrs）
Female10（55.6） 8（44.4）

Treatment tooth

20‒29

30‒39

40‒49

50‒59

60‒69

1（5.6）

3（16.7）4（22.2）3（16.7）7（38.9） 10（50.0） 10（50.0）

Total 18

We used a clinical investigation proforma to obtainbasic information about each patient（age, sex）, theAverage 51.4

Premolar

Molar

Total 20

11 days and 289 days, respectively. The total period of
the investigation was 10 months on average.

intervention site and investigated procedures such as

Before commencing the cavity preparation, local

whether local anesthesia had been administered, if pulp

anesthesia was used in all cases except one. Also, local

protection had been provided, methods for resin coat-anesthesia was used for inlay placement in 10％ of theing, the resin cement used for adhesion, and any clinicalcases.

symptoms before, during, and after the restorative pro-

The resin-coating technique was performed on eachcedure and at the time of latest recall based on thecavity using a single application of HC Ⅱ in eight cases

dental records. Personal information including patient（40％）
and a combination of HC Ⅱ with a flowable resinname and address was excluded from the data forms tocomposite in 12 cases（60％）. No case required direct

maintain anonymity.

pulp capping during the cavity preparation. The resto-

3 ．Data collection and analysis

rations comprised inlays covered under insurance in 14

After the investigation was completed, the evaluation

cases（70％）（metal inlay in 12 cases and a resin com-forms were collected from the dentists. When there

posite inlay in one case）and self-financed inlays in six

was any doubt about the information provided in thecases（30％）（hybrid resin inlay in one case, gold alloy

form, the dentist who completed it was asked directly

inlay in two cases, ceramic inlay in two cases, and a zir-for confirmation and clarification. Thereafter, all data

conia inlay in one case）.

were collated and analyzed.

In 19 cases（95％）, a dual-cure resin cement was used
for the cementation, SA Luting Plus（Kuraray Noritake

Results

Dental, Tokyo, Japan）, G-Luting（GC, Tokyo, Japan）, andother dual-cure resin cements described as listed in the

The results of the investigation are shown in Table 3.

clinical data forms. In one case（5％）, Super-Bond C &In this study, the restorations were performed on pre-B（Sun Medical）, a methyl methacrylate-based resin

molars in 10 cases（50％）（five cases on maxillary pre-cement, was used.

molars and five cases on mandibular premolars）andAfter inlay placement, the clinical outcome wasmolars in 10 cases（50％）（three cases on maxillary

determined to be favorable in 19 cases（95.0％）.

molars and seven cases on mandibular molars）. The

Although discomfort such as pain on drinking cold

average period between cavity preparation and inlay

water was observed before treatment in nine cases

placement and between inlay placement and recall was（45％）, the outcome was determined to be favorable

ODEP Vol. 1, No. 1

40
Table 3

Evaluation outcomes collected from the protocols
Number of cases

Content of survey
Local anesthesia（Percentage of cases）

Cavity preparation

Yes
No
Yes
No

Inlay placement19（95.0）1（5.0）
2（10.0）18（90.0）

Resin-coating procedure

Single application of HC Ⅱ
Combination of HC Ⅱ with a flowable resin8（40.0）12（60.0）

Resin cement

MMA-based resin cement
Dual-cure resin cement1（5.0）
19（95.0）

Restoration category under the national insurance system

Approved
Not approved

14（70.0）6（30.0）Adverse event after placement of the restorationNo
Yes

19（95.0）1（5.0）

after treatment in eight of the nine cases. Of the cases

placement of the restoration in 95％ of cases was deter-where discomfort was experienced before treatment,

mined, which indicated that the resin-coating techniquepain on drinking cold water persisted after treatment

was effective after cavity preparation in vital teeth.

and even at the time of recall occurred in only one case,

The clinical outcomes were favorable in 8/9 cases thatwhich subsequently required a pulpectomy 155 days

originally presented with pain upon cold water stimula-

after the commencement of treatment（Table 3）.

tion before the treatment. The resin-coating technique
was able to protect the prepared dentin surface andDiscussion

isolated it from pain-causing stimuli. However, one casestill experienced pain on cold water exposure at the

The results of this study showed that as cavity

time of recall and finally presented with pulpal symp-

preparation for vital teeth was accompanied by the cut-toms, thereby requiring a pulpectomy. Currently, it isting of intact dentin, local anesthesia needed to be usedchallenging to accurately diagnose the pulp response offor cavity preparation in almost all cases. Some studies

a tooth where the patient experiences discomfort. In

have indicated that damage to the pulp is associatedcases requiring a pulpectomy, it is difficult to diagnosewith the cutting of vital dentin when using an air-tur-

the pulpal condition for an inlay restoration. Under the

23,24）. If protecting the vital tooth struc-current circumstances, in cases with discomfort due totures is prioritized based on the idea of minimally inva-

pulpal sensitivity, the restoration should be performed

sive dentistry, a direct composite restoration should beafter recovery from the discomfort. In cases requiring a

selected for caries treatment. However, when an inlay

restoration, it is crucial to carefully perform the restor-

restoration is inevitably selected as the treatment of

ative procedures after providing the patient with an

choice, the tooth preparation should be carefully per-adequate explanation of the benefits and risks of theformed while using air-water spray from the handpiece

procedure and then obtaining consent.

bine hand piece

and a very light touch preparation to minimize damage

The resin-coating technique may improve the bond-to the pulp. On the other hand, local anesthesia was noting performance of resin cement to dentin for indirectused in one case. The reason was not unclear, but cut-

restorations and also enhance the marginal sealting intact dentin might have been minimal in this case.

between the restoration and the cavity wall26‒28）. This

According to Fusayama’s concept25）, local anesthesia isevaluation used dual‒cure resin cements for luting res-not necessary if cutting is limited only for‘caries-in-

torations in most cases. Such cements are routinely

fected’and‘caries-affected’dentin.

used for the luting of tooth-colored restorations, such as

A favorable outcome free of any discomfort after

resin composite and ceramic inlay/onlay restorations,Dec, 2021

Clinical Assessment of Resin-coating Technique41

because the light-curing of the cement effectively
improves the bond strength of the tooth-colored resto-

Conclusion

rations to the underlying tooth structure. On the other
hand, self-cure resin cements, such as Super-Bond C &B, should ideally be used for metal inlay restorations. It

1．Local anesthesia was used for cavity preparation
in a vital tooth in almost all cases.

was reported, however, that the immediate dentin bond

2．The outcome was favorable after placing the res-strength of Super-Bond C&B was significantly lower

toration with the resin-coating technique using HC Ⅱ

than that of dual-cure resin cements, but after 24 h aalone and HC Ⅱ combined with a flowable resin in 95％

29）high bond strength could be achieved . Further inves-of cases.

tigation of the clinical outcomes of restorations

3．Of the nine patients who experienced discomfort

cemented with various types of resin cement, including

before the procedure, the clinical outcome was favor-

cement removal after restoration placement, is requiredable in eight cases（89％）; however, discomfort per-to expand the current evidence base30,31）.

sisted in one case, which required a pulpectomy（11％）.

Resin coating was performed by using HC Ⅱ alone in

4．These results revealed that the resin-coating

37％ of cases and HC Ⅱ combined with a flowable resin

technique using HC Ⅱ was effective in protecting thein 63％ of cases. As the coating with HC Ⅱ forms a

dentin and pulp after cavity preparation.

thin coating of approximately 5 μm thickness per application, HC Ⅱ does not alter the overall contour of the

Conflict of Interest

prepared surface18,32,33）. Meanwhile, when HC Ⅱ is com-

The authors declare that there are no potential conflictsof interest with regard to the publication of this article.

bined with a flowable resin, any undercuts within thecavity preparation can be easily blocked out by placement of the flowable resin. In this study, interviews
after collecting the evaluation forms confirmed that thecavity morphology had been modified by the additionaluse of a flowable resin. In addition, it has been reported
that HC Ⅱ combined with a flowable resin improved
the adhesion of a resin cement to dentin4,10,11,34,35）. Resincoating using HC Ⅱ alone is suitable for cases requiringhigh-precision preparation methods, such as crown
preparation of abutment teeth for a fixed prosthesis. On
the other hand, resin coating using HC Ⅱ combinedwith a flowable resin is suitable for cases requiringsmooth and stress concentration-free cavity forms suchas resin or ceramic inlays.
The resin-coating technique has been covered by theJapanese national health insurance system for the
preparation of vital teeth since 2019, however, it does
not include inlay and onlay cavities. The results of thisstudy revealed the efficacy of the resin-coating technique on inlays and onlays. It is strongly hoped that theresin-coating technique will be covered within theinsurance system for inlay and onlay cavities in the
future. This study verified the short-term clinical outcomes of the resin-coating technique; however, its longterm clinical outcomes need to be investigated.

References
1）Nikaido T, Tagami J, Yatani H, Ohkubo C, Nihei T, Koizumi H, Maseki T, Nishiyama Y, Takigawa T, Tsubota
Y. Concept and clinical application of the resin-coating
technique for indirect restorations. Dent Mater J 2018;
37: 192‒196.
2）Hu J, Zhu Q. Effect of immediate dentin sealing on preventive treatment for postcementation hypersensitivity.
Int J Prosthodont 2010; 23: 49‒52.
3）Santana VB, de Alexandre RS, Rodrigues JA, Ely C, Reis
AF. Effects of immediate dentin sealing and pulpal pressure on resin cement bond strength and nanoleakage.
Oper Dent 2016; 41: 189‒199.
4）Magne P. Immediate dentin sealing: a fundamental procedure for indirect bonded restorations. J Esthet Res
Dent 2005; 17: 144‒154.
5）Magne P, Kim TH, Cascione D, Donovan TE. Immediate
dentin sealing improves bond strength of indirect restorations. J Prosthet Dent 2005; 94: 511‒519.
6）Ishii N, Maseki T, Nara Y. Bonding state of metal-free
CAD/CAM onlay restoration after cyclic loading withand without immediate dentin sealing. Dent Mater J
2017; 36: 357‒367.
7）Islam M, Takada T, Weerasinghe DS, Uzzaman MA,
Foxton RM, Nikaido T, Tagami J. Effect of resin coating
on adhesion of composite crown restoration. Dent
Mater J 2006; 25: 272‒279.

42

ODEP Vol. 1, No. 1

8）Okuda M, Nikaido T, Maruoka R, Foxton RM, Tagami J.

fracture strength of lithium disilicate inlays and over-

Microtensile bond strengths to cavity floor dentin in
indirect composite restorations using resin coating. J20）Nikaido T, Yoda A, Foxton RM, Tagami J. A resin coat-

Esthet Res Dent 2007; 19: 38‒46.
9）Giannini M, Takagaki T, Bacelar-Sá R, Vermelho PM,
Ambrosano GM, Sadr A, Nikaido T, Tagami J. Influenceof resin coating on bond strength of self-adhesive resin
cements to dentin. Dent Mater J 2015; 34: 822‒827.
10）de Andrade OS, de Goes MF, Montes MA. Marginal
adaptation and microtensile bond strength of composite
indirect restorations bonded to dentin treated withadhesive and low-viscosity composite. Dent Mater 2007;
23: 279‒287.
11）Santos-Daroz CB, Oliveira MT, Góes MF, Nikaido T,
Tagami J, Giannini M. Bond strength of a resin cement
to dentin using the resin coating technique. Braz Oral
Res 2008; 22: 198‒204.
12）van den Breemer CR, Özcan M, Pols MR, Postema AR,
Cune MS, Gresnigt MM. Adhesion of resin cement todentin: effects of adhesive promoters, immediate dentin
sealing strategies, and surface conditioning. Int J Esthet
Dent 2019; 14: 52‒63.
13）Brigagão VC, Barreto LFD, Gonçalves KAS, Amaral M,
Vitti RP, Neves ACC, Silva-Concílio LR. Effect ofinterim cement application on bond strength between
resin cements and dentin: immediate and delayed dentin sealing. J Prosthet Dent 2017; 117: 792‒798.
14）Turkistania A, Sadr A, Shimada Y, Nikaido T, Sumi Y,
Tagami J. Sealing performance of resin cements before
and after thermal cycling: Evaluation by optical coherence tomography. Dent Mater 2014; 30: 993‒1004.
15）Feitosa VP, Medina AD, Puppin-Rontani RM, Correr-Sobrinho L, Sinhoreti MA. Effect of resin coat techniqueon bond strength of indirect restorations after thermal
and load cycling. Bull Tokyo Dent Coll 2010; 51: 111‒
118.
16）van den Breemer CRG, Özcan M, Cune MS, van derGiezen R, Kerdijk W, Gresnigt MMM. Effect of immediate dentine sealing on the fracture strength of lithium
disilicate and multiphase resin composite inlay restorations. J Mech Behav Bio Mater 2017; 72: 102‒109.
17）Ashy LM, Marghalani H, Silikas N. In vitro evaluation of
marginal and internal adaptations of ceramic inlay restorations associated with immediate vs delayed dentin
sealing techniques. Int J Prosthodont 2020; 33: 48‒55.
18）Hironaka NGL, Ubaldini ALM, Sato F, Giannini M,
Terada RSS, Pascotto RC. Influence of immediate dentin
sealing and interim cementation on the adhesion of indirect restorations with dual-polymerizing resin cement. J
Prosthet Dent 2018; 119: 678.e1‒678.e8.
19）Hofsteenge JW, Hogeveen F, Cune MS, Gresnigt MMM.
Effect of immediate dentine sealing on the aging andlays. J Mech Behav Biomed Mater 2020; 110: 103906.
ing technique to achieve minimal intervention in indirect resin composites: A case report. Int Chin J Dent
2003; 3: 62‒68.
21）Kusakabe S, Tsuruta H, Uno M, Burrow MF, Nikaido T.
Clinical assessment of resin-coating technique applied to
exposed dentin after crown preparation. Dent Mater J;
doi: 10. 4012/dmj. 2021‒029〔Epub ahead of print〕.
22）Nikaido T, Nakaoki Y, Ogata M, Foxton R, Tagami J.
The resin-coating technique. Effect of a single-stepbonding system on dentin bond strengths. J Adhes
Dent 2003; 5: 293‒300.
23）Farah RI. Effect of cooling water temperature on thetemperature changes in pulp chamber and at handpiecehead during high-speed tooth preparation. Res Dent
Endod 2018; 44: e3.
24）Oztürk B, U ş ümez A, Oztürk AN, Ozer F. In vitro
assessment of temperature change in the pulp chamber
during cavity preparation. J Prosthet Dent 2004; 91:436‒440.
25）Fusayama T. Clinical guide for removing caries using a
caries-detecting solution. Quintessence Int 1988; 19:397‒401.
26）Takahashi R, Nikaido T, Ariyoshi M, Kitayama S, Sadr
A, Foxton RM, Tagami J. Thin resin coating by dual-application of all-in-one adhesives improves dentin bondstrength of resin cements for indirect restorations. Dent
Mater J 2010; 29: 615‒622.
27）Jayasooriya PR, Pereira PN, Nikaido T, Burrow MF,
Tagami J. The effect of a“resin coating”on the interfacial adaptation of composite inlays. Oper Dent 2003; 28:
28‒35.
28）Murata T, Maseki T, Nara Y. Effect of immediate dentin
sealing applications on bonding of CAD/CAM ceramic
onlay restoration. Dent Mater J 2018; 37: 928‒939.
29）Burrow MF, Nikaido T, Satoh M, Tagami J. Early bonding of resin cements to dentin, ―Effect of bonding environment. Oper Dent 1996; 21: 196‒202.
30）Kitasako Y, Burrow MF, Nikaido T, Tagami J. Effect ofresin-coating technique on dentin tensile bond strengths
over 3 years. J Esthet Restor Dent 2002; 14: 115‒122.
31）Shinagawa J, Inoue G, Nikaido T, Ikeda M, Burrow MF,
Tagami J. Early bond strengths of 4-META/MMATBB resin cements to CAD/CAM resin composite.
Dent Mater J 2019; 38: 28‒32.
32）Jayasooriya PR, Pereira PN, Nikaido T, Tagami J. Efficacy of a resin coating on bond strengths of resin
cement to dentin. J Esthet Restor Dent 2003; 15: 105‒113.
33）Ghiggi PC, Steiger AK, Marcondes ML, Mota EG, Bur-Dec, 2021

Clinical Assessment of Resin-coating Technique43

nett LH Jr, Spohr AM. Does immediate dentin sealing

unfilled/lightly filled adhesive systems. J Esthet Restor

influence the polymerization of impression materials?
Eur J Dent 2014; 8: 366‒372.

Dent 2021; 33: 88‒98.
35）Sultana S, Nikaido T, Matin K, Ogata M, Foxton RM,34）de Carvalho MA, Lazari-Carvalho PC, Polonial IF, deSouza JB, Magne P. Significance of immediate dentin
sealing and flowable resin coating reinforcement for

Tagami J. Effect of resin coating on dentin bonding ofresin cement in class Ⅱ cavities. Dent Mater J 2007; 26:506‒513.

ODEP Vol. 1, No. 1

44

Original Article

ODEP 1（1）：44～51，2021

The Shaping Advantage of M-wire Comparedwith Conventional Nickel-titanium Rotary Instruments
in Heavy Curvature Canals
Akito KASAHARA, Mika TANAKA-SATO,
Noriko MUTOH and Nobuyuki TANI-ISHII
Department of Pulp Biology and Endodontics, Kanagawa Dental University

Abstract
Background: The purpose of this study was to evaluate the shaping advantage of M-wire NiTi ProTaper
NEXT（PTN）compared with a conventional NiTi ProTaper Universal（PTU）file in heavy curvature canals.
The shaping ability was measured by the amount of canal cutting and transportation between the PTN andconventional PTU.
Methods: Root canal shaping by the PTN and PTU was classified into two experimental groups according
to the final tip size, ISO #25 or ISO #40. Eighty-four J-shaped root canals（10°, 20°, 30° apical curvature）in
resin blocks were used.
Results: After adjusting for the level and canal wall side, the mean transportation did not significantly
decrease between the PTN and PTU with ISO #25. Significantly less deviation occurred with the PTN andPTU between 10° and 30° using ISO #40.
Conclusions: The PTN improves file flexibility and enables accurate canal shaping for heavy curvature
canals.
Key words: M-wire, ProTaper NEXT, ProTaper Universal, shaping ability, apical curvature

Corresponding author: Nobuyuki TANI-ISHII, Department of Pulp Biology and Endodontics, Kanagawa Dental University, 82,
Inaoka-cho, Yokosuka, Kanagawa 238‒8580, Japan
TEL: ＋81‒46‒822‒9527, FAX: ＋81‒46‒822‒8856, E-mail: n.ishii@kdu.ac.jp
Received for Publication: June 16, 2021/Accepted for Publication: August 24, 2021
DOI: 10.11471/odep.2021‒006

Dec, 2021

The Shaping Advantage of M-wire NiTi File45

micro-cracks during root canal formation 10,11）. Rootcanal wall displacement after cutting12‒15）, fatigue resis-

Introduction

tance16‒18）, and file breakage resistance19） of PTN have
Rotary dental files with a NiTi alloy containing 56％

been previously studied to determine the shaping abil-（wt）Ni and 44％（wt）Ti have significantly changed

ity. However, no report has objectively compared theroot canal preparation. The NiTi alloy has a cubic crys-

different curvature canal range and shaping ability oftal structure with an austenite phase above the shape

PTN with a PTU in the same shaping system. The pur-recovery temperature1）. The austenite phase changes

pose of this study was to analyze the advantages of theto martensite under force and temperature, but when

PTN and the relationship between the amount of canalthe stress is released, the phase returns to austenite2,3）.

wall removed and canal center transportation com-Consequently, the NiTi alloy has a shape memory effectpared with a PTU.

and super-elasticity without plastic deformation, whichis advantageous for instruments used for root canalMaterials and Methods

preparation.
Given the development of NiTi endodontic instruments targeted at root canals with complex anatomical

1 ．Simulated root canal block and experimental
designconfigurations, the shape memory and super-elastic

Eighty-four simulated J-shaped canal blocks with aproperties of such NiTi instruments have helped pre-curvature of 10°, 20°and 30°（apical diameter/taper: 15/

vent endodontic accidents（such as ledges, root canal06, root canal length 19 mm（working length）, Nissin

4,5）

perforations, apical transportation, and file fractures） .

Plastic Training Block S4-U1, Nissin, Kyoto, Japan）
were

Currently, unique heat and mechanical treatments canused. Three J-shaped canal blocks with different angles

be used to complement the mechanical properties of（S4-U1-10°, 20°, 30°）were cut at 19.0 mm（S4-U1-10°）,

NiTi alloys to improve flexibility and fatigue resistance.

18.5 mm（S4-U1-20°）, and 18.0 mm（S4-U1-30°）from the

In 2007, Sportwire LLC（Langley, OK, USA）devel-root canal orifice at the top of the model by IsoMet

oped a unique heat treatment process with the aim of（Buehler, Tokyo, Japan）and adjusted to allow apical

producing a more flexible NiTi alloy with better fatigue

foramen penetration（Fig. 1）.

6）

resistance than conventional NiTi alloys . Additionally,

We used a PTN file（Dentsply Sirona, Ballaigues,

Berendt developed a NiTi composition in which trace

Switzerland）made by M-wire and a PTU file（Dentsply

elements of less than 1％ were added to produce a NiTi

Sirona）as a control.

alloy7）. Under a heat treatment process, the M-wire

For root canal preparation, there were two final api-

alloy converts to a rhombohedral（R）phase（intermedi-

cal sizes for four groups, ISO #25（1: PTN, 2: PTU）and

ate temperature phase between austenite and marten-ISO #40（3: PTN, 4: PTU）
. Using a J-shaped canal block,site）and the elasticity changes with the change frommeasurements were made by classifying the blocks into

austenite to martensite. Therefore, the flexibility of a

two groups. The apical size and file taper（diameter

8,9）

.

increase amount/mm）of each NiTi file were PTN X1

Additionally, NiTi alloy converted to the R phase is（17/04）, X2（25/06）, X3（30/07）, X4（40/06）, and PTU

conventionally produced by cutting; however, NiTi files

F1（20/04）, F2（25/06）, F3（30/06）, and F4（40/06）,

are produced by a non-cutting rolling step, thereby

which were used to set the final apical size to ISO #25

increasing the file fatigue resistance and fracture resis-

and #40.

NiTi endodontic file can be enhanced by modificationtance.

Three curvature blocks（10°, 20°, 30°; each n＝7）were

The ProTaper NEXT（PTN）file used in this study

used, giving a total of 21 canal blocks for each of theis a next-generation model of ProTaper Universal

ISO #25（1: PTN, 2: PTU）
and ISO #40（3: PTN, 4: PTU）

（PTU）
, which is a modification of the conventional NiTigroups. Therefore, 84 canal blocks were tested. All root

alloy PTU and uses M-wire. PTN was developed as acanal models were attached to the Endo Training

file suitable for severely curved root canals given its

Model Castillo（VDW, Frankfurt, Germany）, and root

increased flexibility and fracture resistance. Comparedcanal preparation was performed in a situation wherewith PTU, the PTN suppresses the occurrence ofthe root canal morphology could not be confirmed.

ODEP Vol. 1, No. 1

46aOrifice

Apical

b

30.2°

20.2°

10.3°

5.8 mmFig. 1c

5.8 mm

5.8 mmJ-shaped canal blocks made of clear resin with diameter of 0.15 mm, taper of 06, and apicalcurvatures of（a）10°,（b）20°, and（c）30°.

2 ．Root canal preparation

reach its working length.

The canals were first scouted with an ISO #15 stain-3）ISO #40 PTN: Using X1, X2, X3, and X4 files inless-steel K-file（Dentsply Sirona）to check patency andsequence, select the PTN mode（300 rpm, 2.0 Ncm）,

to precisely determine the working length（19 mm）.

perform root canal cleaning with 2 ml of distilled water

The canal preparation was performed by a single oper-

when exchanging files, and let the X4 file reach its

ator（specialist of JEA and JCD）using the Ni-Ti（PTN,

working length.

PTU）files in strict accordance with the manufacturer’s
recommendations for each system.

4）ISO #40 PTU: Using S1, S2, F1, F2, F3, and F4
files in sequence, select the PT mode（250 rpm, 3.0‒1.0

Before starting root canal preparation with the PTN

Ncm）, perform root canal cleaning with 2 ml of distilled

and PTU files, flare formation in the upper third of all

water when exchanging files, and save the F4 file afterroot canal orifice openings and smooth file guide path

it reaches its working length.

creation（glide path）up to the root apex were per-3 ．Evaluation of the canal shaping

formed. Flare formation was performed using PTN SX

The amount of resin removed in the outer and innerand PT SX files, which are used for straight line forma-sides of the curved root canal was compared. A stereo-tion only. Furthermore, the glide path was confirmed

scopic microscope（SZX16, Olympus, Tokyo, Japan）andup to #15 using an ISO #15 stainless-steel K filea digital camera（DP71, Olympus）were used for the（Dentsply Sirona）. All NiTi files were washed withmeasurement. The transparent root canal models

purified water using a dedicated engine（X Smart plus,

before and after root canal preparation were superim-

D e n t s p l y S i r o n a）, a n d 1 m l o f E D T A - g e l G l i d e

posed with digital images, and the resulting images（Dentsply Sirona）was applied to the NiTi files.

were transferred to a PC and measured. The measure-

Root canal preparation using PTN and PTU files wasment points were 1, 2, 3, 4, and 5 mm from the apex,

performed by a single dentist with more than 7 years ofand the increase in the root canal width on the outer

experience using NiTi rotary files. The following proce-and inner sides was measured and statistical processing

dure was performed and files were exchanged after

was performed（Fig. 2）.

every five root canal formations.

Root canal center transportation was measured as1）ISO #25 PTN: Using two X1 and X2 files sequen-

the amount of root canal wall cutting in the outer andtially, select the X Smart plus PTN mode（300 rpm, 2.0

inner sides of the curved root canal, and the medianNcm）, perform root canal cleaning with 2 ml of distilleddisplacement of the root canal before and after prepara-water by plastic syringe when exchanging files, andtion was measured. We measured ①the distancereach the working length（19 mm）of the X2 file tobetween shaping and original inner canal wall, ②thecomplete the root canal formation.

distance between shaping and original outer canal wall,2）ISO #25 PTU: Using S1（17/02）, S2（20/04）, F1

and ③the canal width of the shaped canal. The canaland F2 files in sequence, select the PT mode（250 rpm,

center transportation was calculated as the centering

3.0‒1.0 Ncm）
, perform root canal cleaning with 2 ml ofratio using the equation ① －②/③16）. The shaping

distilled water when exchanging files, and let the F2 file

becomes more centered as the centering ratioDec, 2021

The Shaping Advantage of M-wire NiTi File47

with a curvature of 10°to 30°（Table 1）.

Apex
1mm

Inner side

2mm
3mm

2 ．Shaping ability of the final apical size ISO #40
Even with ISO #40 and PTN, the amount of resin
removed from the inner and outer sides was less thanthose in the PTU group for all root canals with a curvature of 10°to 30°（Fig. 3-b, 4-b, 5-b）. For ISO #40 andOuter side
4mm

PTN, the amount of resin removed was significantly
5mm

Fig. 2Pre-instrumentation and post-instrumentation images were superimposed
and the difference between the canal
configuration before and after instrumentation was measured in each of thefive traced levels.

less than that of the PTU group at 1‒5 mm on the innerside with a curvature of 10°（Fig. 3-b）and 1 mm on theouter side at 30°（Fig. 5-b）
. The amount of resin
removed by PTN only increased from 4 to 5 mm on the
inner side compared with the outer side at 20°（Fig.
4-b）
and 30°（Fig. 5-b）
. This tendency was similar to thatof the PTU group. The amount of resin removed by
PTU with ISO #40 significantly increased by 1 mm on
the apex side for all curvatures, and the cutting amounton the inner side increased from 4 to 5 mm on the apexside（p＜0.05）（Fig. 5-b）.

approaches zero.
4 ．Statistical analysis

The change in the root canal transportation was less

The amount of root canal wall cutting and root canalthan 0.05 mm with the 10° to 20° curvatures using PTN,

transportation was measured using statistical analysis

but 0.06 mm at the apex 4 to 5 mm into the root withusing one-way analysis of variance and a multiple com-the 30° curvature. The transportation further signifi-

parison test using the Bonferroni-Dunn test. The level

cantly increased to 0.08 mm（p＜0.05）（Table 2）.

of significance was set at p＜0.05.

In contrast, the root canal transportation of PTU

Resultsside of the apex side at 1 mm in all the root canals with

The root canal shaping ability was evaluated accord-

portation on the inner side increased to 0.07 mm anding to the amount of resin removed and the root canal

0.08 mm at 3 and 4 mm, respectively, even with a cur-transportation. In experimental groups 1 to 4, thevature of 10°, and the inner side displacement of theshowed a displacement of 0.1 mm or more on the outera curvature of 10°to 30°. In the PTU, the canal trans-

amount of resin removed was shown by comparing

root canal of 30° also increased to 0.07 mm and 0.08 mm

PTN and PTU for each curvature（10°, 20°, 30°; Figs. 3‒

at 4 and 5 mm, respectively（Table 2）.

5）. The root canal transportation was classified by PTN
and PTU files（Tables 1 and 2）.

Discussion

1 ．Shaping ability of the final apical size ISO #25
The removal of resin by PTN decreased in both the

The mechanical properties of the NiTi file depend on

inner and outer sides in the ISO #25 curvatures of 10°

the processing method of the file and the surface finish-

to 30° compared with the PTU（Fig. 3-a, 4-a, 5-a）
. Mean-ing method, but it has been reported that the change inwhile, the amount of resin removed by PTU on the

crystal structure by the heat treatment process affects

inner and outer sides was small at 10° to 20° curva-

the flexibility of NiTi alloys9,20‒22）. The PTU used in thistures. The amount of resin removed by PTU increased

study has super-elasticity by the stress-induced trans-slightly at the 30° curvature: the apex side was 1 mm

formation of the martensitic phase with a monoclinicon the outer side, and the apical side was 4 mm and 5

crystal structure to the austenite phase, which is themm on the inner side, although the difference was not

cubic crystal structure of a conventional NiTi alloy23）.

significant（Fig. 5-a）.

Meanwhile, PTN, which is a NiTi alloy processed into

The change in the root canal transportation for both

M-wire by heating and cooling, coexists with austenite,

PTN and PTU was within 0.02 mm for all root canals

martensite, and R phases, and has a monoclinic marten-ODEP Vol. 1, No. 1

48

5

PTU

PTN

outer side（a）measurement point（mm）

measurement point（mm）

PTN

4
3
2
1

inner side
0.3

0.2

0.1

0.0

0.1

0.2

5

outer side

（b）

4

＊

3

＊

2
inner side1

0.3

Total amount of removed resin（Mean±SD（mm））at the inner and outer sides

PTU

0.3

0.2

＊
0.1

0.0

0.1

0.2

0.3

Total amount of removed resin（Mean±SD（mm））at the inner and outer sides

Total amount of removed resin（mm）at the different levels after root canal preparation for
10° apical curvature canals by（a）ISO #25 PTN, ISO #25 PTU, and（b）ISO #40 PTN, ISO#40 PTU＊：Significant difference between PTN and PTU（p＜0.05）

Fig. 3

5

PTU

PTN

outer side（a）measurement point（mm）

measurement point（mm）

PTN

4
3
2
1

inner side
0.3

0.2

0.1

0.0

0.1

0.2

0.3

Total amount of removed resin（Mean±SD（mm））at the inner and outer sides

5

PTU
outer side

（b）

4
3
2
1

inner side
0.3

0.2

＊
0.1

0.0

0.1

0.2

0.3

Total amount of removed resin（Mean±SD（mm））at the inner and outer sides

Total amount of removed resin（mm）at the different levels after root canal preparation for
20° apical curvature canals by（a）ISO #25 PTN, ISO #25 PTU, and（b）ISO #40 PTN, ISO#40 PTU＊：Significant difference between PTN and PTU（p＜0.05）
Fig. 4

site phase and R phase16‒18）. Because the elastic modu-

was analyzed by comparing the cutting characteristics

lus is less than that of the austenite phase, the initialof PTN and PTU under a stress load（formation of aflexibility is improved. Additionally, the M-wire NiTi

curved root canal）. The root canal formation using root

alloy is more flexible than the conventional NiTi alloy

canal models with different apex curvatures of 10° tobecause the austenite phase undergoes a martensitic

30° was analyzed with final apical sizes of ISO #25 andtransformation due to the stress on the file when form-

ISO #40. There was a large difference in the displace-ing a curved root canal. The stress load on the file isment of the root canal wall between the groups. In the19）

reduced, which reduces file corruption .

In this study, the usefulness of the M-wire NiTi alloy

ISO #25 group, there was no significant difference inthe amount of resin removed or canal transportationDec, 2021

The Shaping Advantage of M-wire NiTi File
PTU
outer side

（a）

5

PTN
measurement point（mm）

measurement point（mm）

PTN

4
3
2
inner side

1

0.3

0.2

0.1

0.0

0.1

49

0.2

（b）

5

outer side

4
3
2
1

inner side
0.3

0.3

Total amount of removed resin（Mean±SD（mm））at the inner and outer sides

PTU

0.2

＊
0.1

0.0

0.1

0.2

0.3

Total amount of removed resin（Mean±SD（mm））at the inner and outer sides

Total amount of removed resin（mm）at the different levels after root canal preparation for
30° apical curvature canals by（a）ISO #25 PTN, ISO #25 PTU, and（b）ISO #40 PTN, ISO#40 PTU＊：Significant difference between PTN and PTU（p＜0.05）
Fig. 5

Table 1

Statistical analysis of canal transportation at different levels after root canal preparation using a final apical sizeof ISO #25 in 10°, 20°, and 30° apical curvature models by PTN and PTU.
PTN

apical curvaturelevel（mm）
1
2
3
4
5

PTU

10°

20°

30°

10°

20°

30°

0.00±0.00
0.00±0.00－0.02±0.00－0.02±0.00
0.00±0.00

0.00±0.01－0.01±0.00
0.00±0.01－0.01±0.01
0.00±0.00

0.00±0.01
0.00±0.01
0.00±0.00
0.02＋0.01
0.00±0.01

0.00±0.00－0.01±0.01－0.01±0.01－0.01±0.00
0.01±0.01－0.01±0.01
0.00±0.00
0.00±0.00
0.01±0.00
0.01±0.01－0.02±0.01－0.01±0.01
0.00±0.01
0.02±0.01
0.03±0.02
Mean±SD（mm）

Table 2

Statistical analysis of canal transportation at different levels after root canal preparation using a final apical sizeof ISO #40 in 10°, 20°, and 30° apical curvature models by PTN and PTU.
PTN

apical curvature

10°level（mm）
1 －0.03±0.02＊23

0.00±0.01
0.00±0.01＊

40.00±0.00＊

50.01±0.01

PTU

20°

30°

10°

20°

30°

－0.05±0.01＊－0.05±0.03＊－0.10±0.02＊－0.10±0.03＊－0.13±0.03＊－0.02±0.01－0.02±0.02

0.04±0.01
0.05±0.00

0.07±0.03
0.08±0.03－0.01±0.01－0.01±0.01＊：Significant difference between PTN and PTU（p＜0.05）

0.01±0.02－0.04±0.01

0.08±0.01＊
0.04±0.01

0.03±0.01
0.03±0.01

0.07±0.02＊－0.01±0.01－0.05±0.02

0.01±0.01
0.07±0.02
0.08±0.03
Mean±SD（mm）

ODEP Vol. 1, No. 1

50

associated with the change in the bending angle in PTN
and PTU.

Conclusion

In the ISO #25 group, the amount of resin removed
by PTN and PTU was 1 to 5 mm from the apex of the

The amount of canal wall removed and transported

inner and outer sides for curved root canal angles ofby M-wire ISO #40 PTN was less than that by the con-10°, 20°, and 30°. The measured value was less thanventional ISO #40 PTU in 10° to 30° curvature canals

0.05 mm. Furthermore, the root canal transportationand the original root canal form was maintained,

was less than 0.05 mm and both files could form thealthough there was no significant difference in theroot canal and maintain the original anatomical root

shaping ability for curvature canals by both PTN andcanal morphology. The NiTi file with an ISO #25 tip

PTU of ISO #25. The M-wire PTN improves file flexibil-

showed that the shaping ability of the conventional andity and enables accurate canal shaping for wide andM-wire type NiTi alloy files were similar and useful forheavy curvature canals.

root canal preparation of curved root canal angles
between 10° to 30°.
In the ISO #40 group, the amount of resin removed
significantly changed depending on the root canal curvature between the PTN and PTU files. The PTU
increased the lateral displacement by 1 mm from theapex, and by 4 mm and 5 mm from the apex in theinside of the root canal between the 10° to 30° canals.
The risk of transportation and canal perforation in the
curved root canal was observed. However, the PTN
maintained the original root canal morphology when theamount of resin removed in the inner and outer sides
was 1 to 3 mm and from the apex was 0.1 mm or lessbetween the 10° to 30°canals. For the 30° curvature
canal, the inner side cutting of 4 mm and 5 mm fromthe apex tended to be as high as that of the PTU. The
PTN obtained by converting the NiTi alloy into the Rphase by heat processing improved the root canal followability and suppressed the transport on the outerside of the apex. It is necessary to be careful of accidents, such as strip perforation during root canal formation on the inner side, when using the 30° curvature.
Because the PTN and PTU used in this study are
similar file systems for complete root canal formation
with multiple lines, the flexibility of the M-wire NiTi
alloy is useful for proper root canal formation. Additionally, the M-wire type NiTi alloy single file preparation
system（Reciproc, WaveOne file）completes root canalformation with only one file, the root canal wall displacement is less than that by the PTU, and the root
canal transportation is reduced. Additionally, the PTN
is reported to retain anatomical morphology9,23）. Finally,
the PTN has excellent fatigue resistance and torsion
resistance regardless of the difference in file systems
used for root canal preparation.

Acknowledgements
We thank Ashleigh Cooper, PhD, from Edanz Group（https://en-author-services.edanzgroup.com/）for editing a
draft of this manuscript.
This study was partly supported by a Grant-in-Aid for
Scientific Research from the Ministry of Education, Culture,
Sports, Science and Technology of Japan（B）（No. 20390437）.
The authors have no conflicts of interest related to thisstudy.

References
1）Thompson SA. An overview of nickel-titanium alloys
used in dentistry. Int Endod J 2000; 33: 297‒310.
2）Lee JH, Park JB, Andreason GF, Lakes RS. Thermomechanical study of NiTi alloys. J Biomed Mater Res 1988;
22: 573‒588.
3）Serene TP, Adams JD, Saxena A. Nickel-titanium instruments: Applications Endodontics. St. Louis: Ishiyaku
Euro America Inc.
4）Walia HM, Brantley WA, Gerstein H. An initial investigation of the bending and torsional properties of Nitinolroot canal files. J Endod 1988; 14: 346‒351.
5）Kazemi RB, Stenman E, Spångberg LS. A comparison ofstainless steel and nickel-titanium H-type instruments ofidentical design: torsional and bending tests. Oral Surg
Oral Med Oral Pathol Oral Radiol Endod 2000; 90: 500‒
506.
6）Zinelis S, Darabara M, Takase T, Ogane K, Papadimitriou GD. The effect of thermal treatment on the resistance of nickel-titanium rotary files in cyclic fatigue.
Oral Surg Oral Med Oral Pathol Oral Radiol Endod
2007; 103: 843‒847.
7）Berendet C. Method of preparing Nitinol for use in manufacturing instrument with improved fatigue resistance.

Dec, 2021

The Shaping Advantage of M-wire NiTi File

US patent Application 20070072147 AI.

51

Contemp Dent Pract 2019; 20: 363‒369.

8）Alapati SB, Brantley WA, Iijima M, Clark WAT, Phil D,
Kovarik L, Buie C, Liu J, Johnson WB. Metallurgical16）Özyürek T, Yılmaz K, Uslu G. The effects of autoclave

characterization of a new nickel-itanium wire for rotary
endodontic instruments. J Endod 2009; 35: 1589‒1593.
9）Pereira ESJ, Peixoto IFC, Viana ACD, Oliveira II, Gonzalez BM, Buono VTL, Bahia MGA. Physical and mechanical properties of a thermomechanically treated NiTi
wire used in the manufacture of rotary endodontic
instruments. Int Endod J 2012; 45: 469‒474.
10）De-Deus G, Belladonna FG, Souza EM, Silva EJ, Neves
Ade A, Alves H, Lopes RT, Versiani MA. Micro-computed tomographic assessment on the effect of ProTaper Next and Twisted File Adaptive systems on dentinal cracks. J Endod 2015; 41: 1116‒1119.
11）Saha SG, Vijaywargiya N, Saxena D, Saha MK, Bharadwaj A, Dubey S. Evaluation of the incidence of microcracks caused by Mtwo and ProTaper Next rotary filesystems versus the self-adjusting file: A scanning electron microscopic study. J Conserv Dent 2017; 20: 355‒
359.
12）Hiran-us S, Pimkhaokham S, Sawasdichai J, Ebihara A,
Suda H. Shaping ability of ProTaper NEXT, ProTaper
Universal and iRace files in simulated S-shaped canals.
Aust Endod J 2016; 42: 32‒36.
13）Gagliardi J, Versiani MA, de Sousa-Neto MD, Plazas-Garzon A, Basrani B. Evaluation of the shaping characteristics of ProTaper Gold, ProTaper NEXT, and ProTaper
Universal in curved canals. J Endod 2015; 41: 1718‒1724.
14）Wu H, Peng C, Bai Y, Hu X, Wang L, Li C. Shaping ability of ProTaper Universal, WaveOne and ProTaper
Next in simulated L-shaped and S-shaped root canals.
BMC Oral Health 2015; 15: 27. doi: 10.1186/s12903-0150012.
15）Tavanafar S, Gilani PV, Saleh AM, Schäfer E. Shaping
ability of ProTaper Universal, ProTaper NEXT and
WaveOne Primary in severely curved resin blocks. J

Universal, ProTaper Next, and ProTaper Gold nickel-titanium instruments. Restor Dent Endod 2017; 42: 301‒
308.
17）Topçuoğlu HS, Topçuoğlu G, Akti A, Düzgün S. In vitro
comparison of cyclic fatigue resistance of ProTaper
Next, HyFlex CM, OneShape, and ProTaper Universal
instruments in a canal with a double curvature. J
Endod 2016; 42: 969‒971.
18）Uygun AD, Kol E, Topcu MK, Seckin F, Ersoy I, Tanriver M. Variations in cyclic fatigue resistance among
ProTaper Gold, ProTaper Next and ProTaper Universal
instruments at different levels. Int Endod J 201; 49:494‒499.
19）Fernández-Pazos G, Martín-Biedma B, Varela-Patiño P,
Ruíz-Piñón M, Castelo-Baz P. Fracture and deformationof ProTaper Next instruments after clinical use. J ClinExp Dent 2018; 10: e1091‒e1095.
20）Pereira ES, Gomes RO, Leroy AM, Singh R, Peters O.
Mechanical behavior of M-wire and conventional NiTi
wire used to manufacture rotary endodontic instruments. Dent Mater 2013; 29: e318‒e324.
21）Gutmann JL, Gao Y. Alteration in the inherent metallic
and surface properties of nickel-titanium root canal
instruments to enhance performance, durability andsafety: a focused review. Int Endod J 2012; 45: 113‒128.
22）Braga LC, Silva AC, Buono VT, Bahia MG. Impact ofheat treatments on the fatigue resistance of different
rotary nickel-titanium instruments. J Endod 2014; 40;1494‒1497.
23）Câmara AS, Martins RC, Viana ACD, Leonardo PT,
Buono VTL, Bahia MGA. Flexibility and torsional
strength of ProTaper and ProTaper Universal rotary
instruments assessed by mechanical tests. J Endod
2009; 35: 113‒116.

sterilization on the cyclic fatigue resistance of ProTaperODEP Vol. 1, No. 1

52

Original Article

ODEP 1（1）：52〜58，2021

Influence of Surface Moisture on the Bond Strengthof a Universal Adhesive in the Etch-and-rinse Mode
Runa SUGIMURA, Akimasa TSUJIMOTO＊, Kei IWASE,
Shun KATSUKI, Ahmad ALKHAZALEH＊, Olajide OBE＊,
Toshiki TAKAMIZAWA and Masashi MIYAZAKI
Department of Operative Dentistry, Nihon University School of Dentistry＊

Department of Operative Dentistry, University of Iowa College of Dentistry

Abstract
Purpose: This study aimed to investigate the effect of surface moisture on the enamel and dentin bonding
performances of a universal adhesive in the etch-and-rinse mode based on shear bond strength（SBS）tests.
Methods: Enamel and dentin surfaces were prepared（moist and dry conditions）using bovine mandibular
incisors and treated with the universal adhesive Clearfil Universal Bond Quick（CU）using the etch-and-rinsemode. Resin composite was condensed into a plastic mold clamped to a fixture against the adherend surfaces
and light-cured for 40 s. The bonded specimens（n＝15）were either stored in distilled water at 37℃ for 24 hor underwent 10,000 thermal cycles between 5℃ and 55℃ before the SBS tests. The universal adhesiveinterfaces in the etch-and-rinse mode were observed via scanning electron microscopy（SEM）.
Results: The CUʼs enamel and dentin bond strengths were not influenced by the surface moisture, regardless of the storage condition. Based on the SEM observations, surface moisture did not influence the widthsof the adhesive and hybrid layers in the resin-dentin interfaces. However, the lengths of the resin tags in the
moist group were longer than those in the dry group.
Conclusion: These findings indicated that the surface moisture content of the enamel and dentin might notsignificantly influence CU bonding performance regardless of the storage conditions. The presence of a newhydrophilic amide monomer in this universal adhesive might be a key factor that influences its bonding performances with enamel and dentin.
Key words: universal adhesive, surface moisture, etch-and-rinse mode

Corresponding author: Toshiki TAKAMIZAWA, Department of Operative Dentistry, Nihon University School of Dentistry,
1 8 13, Kanda-Surugadai, Chiyoda-ku, Tokyo 101 8310, Japan
TEL: ＋81 3 3219 8141, FAX: ＋81 3 3219 8347, E-mail: takamizawa.toshiki@nihon-u.ac.jp
Received for Publication: July 27, 2021/Accepted for Publication: September 21, 2021
DOI: 10.11471/odep.2021 007

Dec, 2021

Influence of Surface Moisture on Bond Strength of Universal Adhesive

Introduction

53

Materials and Methods

Universal adhesives, in both etch-and-rinse and self-

1 ．Study materials

etch modes, have increased in popularity due to their

The universal adhesive used in this study（Clearfil

versatility and relative ease of application1）. Universal

Universal Bond Quick ER; CU）was obtained from Kur-

adhesives can be used as pretreatment agents for resin

aray Noritake Dental, Tokyo, Japan. Ultra-Etch（Ultra-

luting cements2）; they can bond to various substrates

dent Products, South Jordan, UT, USA）
, a 35％ phos-

without surface pretreatment

3）and can be used with

reduced application time4）.

phoric acid solution, was used as the etching agent, andClearfil AP-X（Kuraray Noritake Dental）was used as

A recent systematic review on the laboratory bondthe resin composite to fabricate the specimens（Table 1）.

strength of universal adhesives reported that the dentin

2 ．Specimen preparationbond strength of universal adhesives was not affected

The shear bond strengths of CU to enamel and den-by the bonding strategy employed. In contrast, thetin in the etch-and-rinse mode were measured using theenamel bond strength was lower in the self-etching

notched-edge test specified by ISO 29022 11）. Bovine

mode than in the etch-and-rinse mode1）. In addition,

mandibular incisors were extracted from 2- to 3-year-

previous studies of universal adhesives demonstrated

old cattle. The roots were cut using a slow-speed saw

that the use of an etch-and-rinse mode for dentin bond-equipped with a diamond-impregnated disk（Isomet,

ing yields a bond strength equal to or greater than a

Buehler, Lake Bluff, IL, USA）
, and the pulps were

5‒7）

. These results suggested that univer-removed. Each tooth was then mounted in self-curing

sal adhesives might be more suitable for use in theacrylic resin（Tray Resin Ⅱ, Shofu, Kyoto, Japan）and

etch-and-rinse mode than in the self-etch mode in someplaced underwater to limit the temperature rise caused

cases. However, there is no consensus on the optimalby the exothermic polymerization of the resin. The den-conditions for using universal adhesives in the etch-and-tin bonding surfaces were ground flat using a grind-

rinse mode; the ideal surface moisture condition for

er-polisher（EcoMet 4, Buehler）with P120 and P400

applying these adhesives has not been determined sosilicon carbide（SiC）papers（Struers, Cleveland, OH,

far. The influence of surface moisture on the bondingUSA）under running water. The surfaces were thenperformance of dental adhesive systems has long been

washed with an air-water spray and air-dried using a

discussed. A higher water content or over-drying of thethree-way syringe.

self-etch modedemineralized dentin can degrade the quality of the8）

The prepared enamel and dentin adherent surfaces

adhesive layer and reduce the bond strength . Previous

were etched with 35％ phosphoric acid for 15 s before

studies have reported that the dentin bond strength ofapplying the adhesive. Subsequently, they were divided

universal adhesives in the etch-and-rinse mode is notinto two groups based on the level of moisture: moist

affected by surface moisture9,10）. Nonetheless, the effectgroup, where 2.5μl of distilled water was applied to anof surface moisture on the bonding of universal adhe-area（diameter, 4 mm）and blot dried; and dry group,

sives to enamel and dentin in the etch-and-rinse mode

where 2.5μl distilled water was applied to an area ofneeds to be clarified.

similar diameter and air-dried for 5 s using a dental

This study aimed to investigate the influence of sur-three-way syringe（distance, 5 cm above the surface; airface moisture on the bonding performance of a univer-pressure, 0.25 MPa）
. The specimens were prepared

sal adhesive in the etch-and-rinse mode. The nullunder ambient laboratory conditions at 23℃±2℃ andhypotheses tested were as follows: 1）the surface mois-

50％±10％ relative humidity.

ture in phosphoric acid-etched enamel and dentin does

3 ．Shear bond strength testing

not influence the bond durability of the universal adhe-

The prepared enamel and dentin surfaces from thesive, and 2）the surface moisture does not affect thetwo groups were treated with the universal adhesiveinterfacial morphology of the universal adhesive in the

per the manufacturer’s instructions. After applying the

etch-and-rinse mode.

adhesive to the tooth surface, a Bonding Mold Insert（2.38 mm in internal diameter and 2.5 mm in height,

ODEP Vol. 1, No. 1

54
Table 1
Material

Type

Materials used in this study
Main components

Manufacturer（Lot No.）（Code）

Clearfil Universal
Bond Quick ER（9T0050）

Adhesive（CU）

Ultra‒Etch（A124）

Etching agent

35％ phosphoric acid, glycol, cobalt aluminate blue spinel

Clearfil AP‒X
（380094）

Resin composite

bis‒GMA, TEGDMA, silanated barium glass filler, silaKuraray Noritake Dental,
nated silica filler, silanated colloidal silica, CQ, pigments,
Tokyo, Japanothersbis‒GMA, MDP, HEMA, hydrophilic amide monomer,
Kuraray Noritake Dental,
filler, ethanol, water, NaF, photo initiators, chemicalTokyo, Japanpolymerizer, accelerator, silane coupling agent, others
Ultradent Products, South
Jordan, UT, USA

Ultradent Products）was clamped to a fixture against

and the tooth bonding sites. The failure modes were

the adherend surfaces and filled with resin compositeclassified as follows: adhesive failure at the interface;

using a condensing instrument. The resin composite

cohesive failure in the resin composite; cohesive failure

was light-cured for 40 s at a standardized distance of 1

in the enamel or dentin; and mixed failure.

mm using a quartz-tungsten halogen（QTH）curing unit

5 ．Scanning electron microscopy obser vation（Optilux 501, Kerr, Orange, CA, USA）
. The light inten-

Representative micrographs of the resin-enamel andsity（＞600 mW/cm2）of the QTH curing unit was mon-resin-dentin interfaces were obtained using field-emis-

itored using a curing radiometer（Model 100, Kerr）. The

sion scanning electron microscopy（SEM; ERA 8800FE,

plastic mold was removed, and the bonded specimens

Elionix, Tokyo, Japan）. Acid-base treatment was per-were stored in distilled water at 37℃ for 24 h. The

formed to visualize the resin tags and assess their

specimens were randomly allocated to two subgroups

length to dentinal tubules. The enamel and dentin sur-

（n＝15 each）as follows: those that did not undergo

faces were prepared（as described earlier）, and thethermal cycling（no thermal cycling; 24 h）
and those thatbonded specimens were stored in distilled water atunderwent 10,000 thermal cycles（10,000 TC at 5℃ and

37℃ for 24 h. They were embedded in epoxy resin

55℃）. Thermal cycling was conducted using a thermal（Epon 812, Nisshin EM, Tokyo, Japan）and stored atshock tester（TTS-1, Thomas Kagaku, Tokyo, Japan）,

37℃ for an additional 24 h. Subsequently, the specimens

and each cycle involved incubation in each water bathwere sectioned at the center, and the surfaces of thefor 30 s with a transfer time of 5 s.
The shear bond strength（SBS）measurements were

cut halves were polished using a grinder-polisher（#4,000-grit SiC paper）. Finally, the surfaces were pol-performed using a universal testing machine（5500R,

ished with a soft cloth using 0.25μm-grit diamond paste

Instron, Norwood, MA, USA）equipped with Test Base

（DP-Paste; Struers, Ballerup, Denmark）and ultrasoni-

Clamp（Ultradent Products）at a crosshead speed of 1.0

cally cleaned for 3 min. The polished surfaces of speci-

mm/min. Crosshead Assembly（Ultradent Products）

mens with the resin-dentin with acid-base treatment

with a semicircular notch（diameter, 2.38 mm）was usedinterfaces were etched with HCl solution（6 mol/l）forfor the measurements. The SBS values（MPa）were

25 s and deproteinized by immersion in 6％ NaOCl

calculated by dividing the peak load at failure by thesolution for 3 min. The SEM specimens were dehy-

bonding area.

drated by immersing them in ascending concentrations

4 ．Failure mode analysis

of aqueous tert-butanol（50％ for 20 min, 75％ for 20 min,After the SBS tests, a single experienced individual

95％ for 20 min, and 100％ for 2 h）and then transfer-

evaluated the bond failure sites via optical microscopy

ring them to a freeze dryer（Model ID-3, Elionix）for 3（SZH-131, Olympus, Tokyo, Japan; magnification, 20×）.

min. In order to enhance the visibility of the layers on

The failure mode was assessed based on the percent-the adhesive interfaces, the specimens’ polished sur-age of surface area（adhesive, resin, or substrate）

faces were etched for 30 s using an argon ion-beamobserved on both the bonded resin composite cylinders（Type EIS-200ER, Elionix）, which was directed perpen-Dec, 2021

Influence of Surface Moisture on Bond Strength of Universal Adhesive
Table 2

55

Shear bond strength（MPa）and standard deviations（in parentheses）of theuniversal adhesive with their failure mode

Substrate

Surfacewetness

EnamelDentin24 h

10,000 TC

Bond strength（MPa）

Failure mode

Bond strength（MPa）

Failure mode

Moist

33.4（3.9）a［100/0/0/0］［100/0/0/0］

Dry

34.8（5.0）［100/0/0/0］a35.6（4.7）

a

Moist

b

21.2（4.5）［93/0/7/0］

Dry

22.5（4.8）b［86/0/14/0］a36.9（5.3）［100/0/0/0］b［100/0/0/0］

23.1（4.7）
24.4（5.1）b［100/0/0/0］

Same small letters indicate no significant differences（p＞0.05）.
Percentage of failure mode
［adhesive failure/cohesive failure in resin/cohesive failure
in enamel or dentin/mixed failure］. There were no significant differences in failuremode with surface wetness or substrate.

dicular to the surface at an accelerating voltage of 1.0

exact tests did not reveal statistically significant differ-

kV and an ion current density of 0.4 mA/cm2. The sur-ences in failure modes based on the surface moisturefaces were then coated with a thin film of gold in a vac-conditions or the substrate.

uum evaporator（Quick Coater Type SC-701, Sanyu

3 ．SEM obser vation of bonding interface

Electron, Tokyo, Japan）and observed using field-emis-

Figure 1 shows representative SEM micrographs ofsion SEM（operating voltage, 10 kV）. The approximate

the control and etching groups. SEM observations oflengths of the resin tags were measured on the images

the resin-enamel and resin-dentin interfaces showedof the acid-base-treated resin-dentin interfaces.

excellent adaptation, regardless of surface moisture.

6 ．Statistical analysis

The thickness of the adhesive layers in the resin-

The SBS data were analyzed using a two-way analy-

enamel and resin-dentin interfaces ranged from 5 to 10

sis of variance followed by Tukey’s post hoc honestly

μm; a hybrid layer（thickness, approximately 2μm）was

significant difference test. Fisher’s exact test was usedvisible in the resin-dentin interfaces. However, the sur-for statistical analysis of the failure mode after SBS

face moisture did not influence the widths of the adhe-testing. The significance level in this study was set atsive and hybrid layers. In the SEM observations of thep＜0.05. The statistical analysis was conducted using aresin-dentin interfaces treated with an acid-base, thecommercial statistical software package（SPSS Statis-

resin tags were identified and concentrated in the den-tics, IBM, Armonk, NY, USA）.

tinal tubules. The length of the resin tag was dependent
on the surface moisture condition of the dentin; theResults

length in the moist group was longer（40‒60μm）thanthat in the dry group（30‒50μm）.

1 ．SBS
Table 2 shows the results for the effect of surface

Discussion

moisture on the SBS of CU in the etch-and-rinse mode.
The SBS to the enamel was not influenced by the sur-Clinicians try to control the surface moisture of theface moisture, regardless of the storage condition. Fur-

substrate after phosphoric acid etching when using uni-

thermore, no significant difference（p＞0.05）in the SBS

versal adhesives in the etch-and-rinse mode. However,of the universal adhesive to dentin between the moist

moisture control is dependent on the individual’s inter-and dry groups was noted, regardless of the storage

pretation of the most appropriate surface moisture con-condition.

dition for the procedure. This study investigated the

2 ．Failure mode

influence of surface moisture on the bonding strengths

The results of the failure mode analysis for SBS afterof a universal adhesive to both enamel and dentin in the

24 h and 10,000 TC are shown in Table 2. Fisher’s

etch-and-rinse mode.

ODEP Vol. 1, No. 1

56

R

R

R
HL

A

RT

A
E
Moist-Enamel

D

R

R

A

A

E

Dry-Enamel

D

Moist-Dentin-Acid-Base

Moist-Dentin

R

HL

RT
Dry-Dentin

Dry-Dentin-Acid-Base

Representative scanning electron microscopy images of resin-enamel, resin-dentin, and resin-dentin＋acid-basetreated interfaces
The left upper and lower images indicate representative resin-enamel interfaces. The upper and lower images in the
middle indicate representative resin-dentin interfaces. The main images are at ×5,000 magnification. The smaller white
rectangle indicates the location in the main images at ×20,000. The right upper and lower images indicate representative
resin-dentin interfaces with acid-base treatment. The main images are at ×1,000 magnification. The smaller white rectangle indicates the location in the main images at ×5,000. A: adhesive; Acid-Base: acid-base treatment; D: dentin; E: enamel;
HL: hybrid layer; R: resin; RT: resin tags.
Fig. 1

The results of the present study suggested that theOn the other hand, the inorganic apatite crystals insurface moisture of adherent surfaces did not affect thethe smear layer, superficial layer, and subsurface of thebond durability of CU in the etch-and-rinse mode.

phosphoric acid-etched dentin are demineralized by

Therefore, the first null hypothesis that“surface mois-

etching and replaced by water. The intrinsic water-ture does not affect the bond durability of universal

bound hydroxyl groups surround the collagen fibrils17）.

adhesives in the etch-and-rinse method”was not

The water must surround the collagen fibrils to main-

rejected. These results were consistent with those of atain the rigidity of the spaces between the fibrils,

previous study12）, even when using the current univer-increase the adhesive penetration into the collagen net-

sal adhesive. Phosphoric acid etching creates awork, and facilitate the interaction between the adhe-13）

and increases the surface free

sive and the collagen. When the dentin surface becomesenergy of enamel14）. In contrast, in the case of the den-excessively dry, the collagen network collapses andtin, the surface becomes hydrophobic, and the surface

shrinks, trapping water within the dentin tubules,

free energy is decreased 15）. Previous studies haveresulting in poor interaction with the adhesive andreported that the surface of the enamel etched with

reduced adhesive penetration18）. In contrast, when thephosphoric acid is more“friendly”toward water thansurface of the etched dentin is over-wet, the collagenthat of the dentin because the water is tightly trapped

network swells, and the interfibrillar space decreases,

onto the surface owing to its structure and high surface

resulting in poor penetration of the adhesive into themicro-porous surface

16）

free-energy . This makes it difficult for the water todentinal tubules19）. Furthermore, excessive moisture on

be removed by air-drying and the small difference inthe substrate causes phase separation of the adhesive20）

water content between the wet and dry enamel sur-and poor polymerization21）, leading to hydrolysis andfaces when compared to those of dentin.

degradation of the adhesive layer. Thus, moisture con-

Dec, 2021

Influence of Surface Moisture on Bond Strength of Universal Adhesive

57

trol on the surface of demineralized dentin is an import-

surface wetting conditions. Therefore, the new hydro-

ant factor determining the adhesive effect of the adhe-philic amide monomer might reduce the effect of sur-22）

sive in the etch-and-rinse mode .

face moisture on the bond durability of CU. By captur-

The effect of the surface moisture condition was visi-

ing the moisture from the substrate, the universal adhe-ble in the SEM images of the resin tags in the acid-sive makes it more prone to demineralization andbase-treated resin-dentin interface. Moreover, there

increases the chemical reactions between the adhesive

were no differences in the widths of the adhesive andand the substrate. Excess moisture in the adhesive will

hybrid layers between the moist and dry groups at theresult in insufficient polymerization and a low-quality

resin-enamel or resin-dentin interfaces. The length ofadhesive layer. In contrast, insufficient moisture in thethe resin tags in the acid-base-treated resin-dentin

adhesive will result in weak demineralization andinterfaces depended on the surface moisture conditions;poorer chemical reactions. Therefore, it is important to

the resin tags in the moisture group were longer than

optimize the moisture content of the adhesive before

those in the dry group. However, no difference in theapplying it to the tooth substrate.

thickness of the hybrid layer was noted. Therefore, theNowadays, the principle of development differs

second null hypothesis,“the interfacial morphology ofamong manufacturers, even for adhesives that belong to

CU in the etch-and-rinse mode does not differ depend-

the same category. Therefore, the effect of surface

ing on the surface moisture,”was partially rejected.

moisture on bond durability in the etch-and-rinse mode

In the present study, no relationship between thecan be overcome by adding special ingredients to thelength of the resin tags and dentin bond durability ofadhesive and optimizing the moisture content. How-

CU was observed. A previous study reported no cor-ever, there is little information about the influence ofrelation between the length of the resin tag and the

moisture condition on the bond performance of univer-adhesive strength of a two-step etch-and-rinse adhe-sal adhesives in the self-etch mode. Further research is

23）

sive , but a strong correlation was observed with theneeded to investigate the influence of moisture condi-

thickness of the hybrid layer. In another study, thetion on bond performance when using universal adhe-lengths of the resin tags in the case of the universal

sives in the self-etch mode.

adhesives were significantly longer than those of thetwo-step etch-and-rinse adhesives; no relationship wasConclusion

observed between the length of the resin tag and thedentin bond strength22）. Alternatively, according to

1．The enamel and dentin bond strengths of CU

Sugimura et al., there was an association between thewere not influenced by the surface moisture, regardlesslength of the resin tags and the dentin bond durabilityof the storage condition.

of universal adhesives, and it was material depen24）

dent .

2．SEM observations of the resin-enamel and resin-dentin interfaces showed excellent adaptation,

However, the length of the resin tags and the thick-regardless of the surface moisture. On the other hand,ness of the hybrid layer alone cannot fully explain thethe lengths of the resin tags in the acid-base-treatedresults obtained in the present study because the effectresin-dentin interfaces differed according to the surfaceof surface moisture on the bond durability to dentin

moisture conditions of the dentin.

might be dependent on the material used. The most
likely reason for the difference between the results of

Acknowledgments

this study and those of previous studies might be theThis work was supported in part by Grants-in-Aid for Scientific Research, No. 19K10158, and 21K09900 from theJapan Society for the Promotion of Science. This project wasalso supported in part by the Sato Fund（2021）and by agrant from the Dental Research Center of the Nihon University School of Dentistry（2021）, Japan.

composition of the adhesive used9,10）. CU contains a newhydrophilic amide monomer, which is considered to be
an important technical factor of this adhesive. It is used
as an alternative to 2-hydroxyethyl methacrylate（HEMA）. It exhibits a higher degree of polymerization
than HEMA, thereby reducing water absorption, which
is important to ensure adhesiveness under different

ODEP Vol. 1, No. 1

58
Conflicts of interest
The authors declare no conflict of interest related to thispaper.

References
1）Cuevas-Suárez CE, da Rosa WLO, Lund RG, da Silva AF,
Piva E. Bonding performance of universal adhesives: An
updated systematic review and meta-analysis. J Adhes
Dent 2019; 21: 7‒26.
2）Tsujimoto A, Barkmeier WW, Takamizawa T, Watanabe H, Johnson WW, Latta MA, Miyazaki M. Simulated localized wear of resin luting cements for universal adhesive systems with different curing mode. J OralSci 2018; 60: 29‒36.
3）Siqueira F, Cardenas AM, Gutierrez MF, Malaquias P,
Hass V, Reis A, Loguercio AD, Perdigão J. Laboratory
performance of universal adhesive systems for luting
CAD/CAM restorative materials. J Adhes Dent 2016;
18: 331‒340.
4）Ahmed MH, Yoshihara K, Mercelis B, Van Landuyt K,
Peumans M, Van Meerbeek B. Quick bonding using auniversal adhesive. Clin Oral Investig 2020; 24: 2837‒2851.
5）Nagarkar S, Theis-Mahon N, Perdigão J. Universal dental adhesives: Current status, laboratory testing, andclinical performance. J Biomed Mater Res B Appl Biomater 2019; 107: 2121‒2131.
6）Takamizawa T, Barkmeier WW, Tsujimoto A, Berry
TP, Watanabe H, Erickson RL, Latta MA, Miyazaki M.
Influence of different etching modes on bond strengthand fatigue strength to dentin using universal adhesive
systems. Dent Mater 2016; 32: e9‒e21.
7）Takamizawa T, Barkmeier WW, Tsujimoto A, Suzuki T,
Scheidel DD, Erickson RL, Latta MA, Miyazaki M.
Influence of different pre-etching times on fatigue
strength of self-etch adhesives to dentin. Eur J Oral Sci
2016; 124: 210‒218.
8）Yamamoto K, Suzuki K, Suwa S, Miyaji H, Hirose Y,
Inoue M. Effects of surface wetness of etched dentin on
bonding durability of a total-etch adhesive system: Comparison of conventional and dumbbell-shaped specimens.
Dent Mater J 2005; 24: 187‒194.
9）Perdigão J, Sezinando A, Monteiro PC. Laboratory bonding ability of a multi-purpose dentin adhesive. Am J
Dent 2012; 25: 153‒158.
10）Lenzi TL, Raggio DP, Soares FZM, Rocha Rde O. Bonding performance of a multimode adhesive to artificially-induced caries-affected primary dentin. J Adhes Dent
2015; 17; 125‒131.
11）International Organization for Standardization. Dentistry-Adhesive-Notched-edge shear bond strength test,

ISO 29022 TR: 2013, Geneva.
12）Barkmeier WW, Erickson RL, Latta MA. Fatigue limitsof enamel bonds with moist and dry techniques. Dent
Mater 2009; 25: 1527‒1531.
13）Zhu JJ, Tang AT, Matinlinna JP, Hägg U. Acid etchingof human enamel in clinical applications: A systematic
review. J Prosthet Dent 2014; 112: 122‒135.
14）Tsujimoto A, Barkmeier WW, Takamizawa T, Watanabe
H, Johnson WW, Latta MA, Miyazaki M. Influence ofduration of phosphoric acid pre-etching on bond durability of universal adhesives and surface free-energy characteristics of enamel. Eur J Oral Sci 2016; 124: 377‒386.
15）Yamauchi K, Tsujimoto A, Jurado CA, Shimatani Y,
Nagura Y, Takamizawa T, Barkmeier WW, Latta MA,
Miyazaki M. Etch-and-rinse vs self-etch mode for dentin
bonding effectiveness of universal adhesives. J Oral Sci
2019; 61: 549‒553.
16）Walls AW, Lee J, McCabe JF. The bonding of composite
resin to moist enamel. Br Dent J 2001; 191: 148‒150.
17）Pashley DH, Tay FR, Breschi L, Tjäderhane L, Carvalho
RM, Carrilho M, Tezvergil-Mutluay A. State of the art
etch-and-rinse adhesives. Dent Mater 2011; 27: 1‒16.
18）Wang Y, Spencer P. Hybridization efficiency of the adhesive/dentin interface with wet bonding. J Dent Res
2003; 82: 141‒145.
19）Mohan B, Kandaswamy D. A confocal microscopic evaluation of resin-dentin interface using adhesive systems withthree different solvents bonded to dry and moist dentinan in vitro study. Quintessence Int 2005; 36: 511‒521.
20）Tay FR, Gwinnett AJ, Wei SH. The overwet phenomenon: An optical, micromorphological study of surface
moisture in the acid-conditioned, resin-dentin interface.
Am J Dent 1996; 9: 43‒48.
21）Toledano M, Cabello I, Yamauti M, Giannini M, Aguilera
FS, Osorio E, Osorio R. Resistance to degradation of resin-dentin bonds produced by one-step self-etch adhesives. Microsc Microanal 2012; 18: 1480‒1493.
22）Iwami Y, Yamamoto H, Kawai K, Ebisu S. Effect ofenamel and dentin surface wetness on shear bondstrength of composites. J Prosthet Dent 1998; 80: 20‒26.
23）Anchieta RB, Oliveira FG, Sundfeld RH, Rahal V, Machado LS, Alexandre RS, Sundefeld ML, Rocha EP. Analysis of hybrid layer thickness, resin tag length and theircorrelation with microtensile bond strength using a total
etch adhesive to intact dentin. Acta Odontol Latinoam
2011; 24: 272‒278.
24）Sugimura R, Tsujimoto A, Hosoya Y, Fischer NG, Barkmeier WW, Takamizawa T, Latta MA, Miyazaki M.
Surface moisture influence on etch-and-rinse universal
adhesive bonding. Am J Dent 2019; 32: 33‒38.

Dec, 2021

59

Original Article

ODEP 1（1）：59～68，2021

Removal of Biofilm from Titanium Surfaces Using 400-kHz UltrasoundTakashi TAKIGUCHI, Junki YAMADA,
Masanori SATO＊ and Matsuo YAMAMOTO
Department of Periodontology, Showa University School of Dentistry＊

Honda Electronics Co., Ltd.

Abstract
Purpose: Peri-implantitis is a microbial infectious disease that is similar to periodontal disease. Although
one of the most important procedures for managing peri-implantitis is to remove dental biofilm from titanium surfaces, this is less than adequate when the procedure is conducted using conventional dental devices.
Ultrasound in the MHz range generates vibration in water, thereby accelerating water molecules throughwhich debris on the surface of an object can be removed. The objective of this study was to investigate theeffectiveness of ultrasonic water flow in removing plaque from titanium specimens.
Methods: Sound pressure was measured by the beam pattern distribution and a hydrophone. The optimum applied power and irradiation conditions were determined based on the relationship between artificial
plaque removal effect and sound pressure. Plaque biofilms were formed over a period of 72 h on the surfacesof titanium specimens attached to custom-made stents fixed to the buccal surfaces of the molar and premolar teeth of volunteers. In this experiment, the titanium surfaces were exposed to an ultrasonic water flow of
400 kHz, with a sound pressure of 0.6 MPa, for 180 s. After cleaning by the ultrasonic water flow, the titanium surfaces were examined by a scanning electron microscope. For each specimen, the residual plaquebiofilm areas were evaluated as percentages of the surface scanned.
Results: The sound pressure measurements were confirmed to be reasonably accurate based on the calculated results of the beam pattern distribution. The artificial plaque was removed in a sound pressure-dependent manner. The total proportions of residual plaque biofilm were 22.8％（median）and 7.05％（median）on
mirror-finished and rough surfaces, respectively. The difference in residual plaque biofilm compared to thecondition before treatment on both the mirror-finished and rough surfaces was judged to be significant at＊

p＜0.05. Furthermore, the ultrasonic water flow completely destroyed the bacterial chains on the titanium

specimens.
Conclusions: This study demonstrated that the 400 - kHz, 0.6 - MPa sound pressure ultrasonic water flowcleaning device can effectively remove plaque biofilm from titanium surfaces.
Key words: ultrasonics, vibrational acceleration, sound pressure, biofilm

Corresponding author: Matsuo YAMAMOTO, Department of Periodontology, Showa University School of Dentistry, 2‒1‒1,
Kitasenzoku, Ohta-ku, Tokyo 145‒8515, Japan
TEL: ＋81‒3‒5498‒1995, FAX: ＋81‒3‒3787‒7886, E-mail: yamamoto-m@dent.showa-u.ac.jp
Received for Publication: July 5, 2021/Accepted for Publication: September 30, 2021
DOI: 10.11471/odep.2021‒008

ODEP Vol. 1, No. 1

60

Recently, we focused on developing a new cleaning system operating at a midrange frequency, in which 400 -Introduction

kHz ultrasound was superposed on a water flow from a
Peri-implantitis is a microbial infectious disease that

nozzle. The ultrasonic water flow（UWF）worked as ais similar to periodontal disease. One of the most

medium not only for propagating ultrasound but also

important procedures for managing peri-implantitis is

for transporting debris detached by the ultrasonic

the removal of dental biofilm from titanium surfaces.

action17）.

Biofilm consists of microbes enclosed within extracellu-

The present study aimed to explore whether it islar polymeric substances such as polysaccharides, pro-possible to use the developed UWF system to remove

teins, and nucleic acids. Biofilm formation can be

biofilm from titanium surfaces.

observed on the surfaces of medical devices such as
venous catheters, heart valves, coronary stents, andMaterials and Methods

dental implants1,2）. The biofilm formed around dental
implants follows a similar pattern of microbial coloniza-

1 ．Ultrasonic water flowtion as that formed on natural teeth3）. Microbiological

UWF is a non-contact high-frequency ultrasonicevidence of biofilm-related peri-implant infection has

cleaning system that removes dental biofilm from thebeen reported in the literature4,5）. Peri-implant mucosi-

minute structures of implants without cavitation or con-

tis and peri-implantitis were reported as new dental

tact damage.

diseases affecting implants6,7）. Although thorough bacte-

Figure 1 depicts a schematic of the experimental

rial reduction is necessary to treat peri-implant dis-

setup of the UWF system. The washing fluid is sup-eases, it is difficult to eliminate bacteria from a dental

plied to the device from a water tank, in which ultra-

implant surface using conventional tools such as tooth

sonic waves（frequency: 400 kHz; input power: 0‒25 W）

8,9）

brushes and ultrasonic scalers, which is because theare propagated. The diameter of the ultrasonic transducer is 30 mm. The nozzle’s outlet diameter is 6 mm.

implant’s roughness is in the order of micrometers.
Ultrasonic scalers have an advantage with respect to

The vibrator case is tapered toward an outflow port,

the removal of plaque and calculus, but they also carryand the washing fluid through which the ultrasonic

10,11）

the risk of scratching the surface of dental implants.

Therefore, it is necessary to develop a device for clean-waves propagate impinges upon the specimen.
The UWF device was mounted on an arm that could
be adjusted along three axes to ensure reproducibility.

ing dental plaque from rough surfaces.
Ultrasonic cleaning is an efficient method to remove

In the present experiment, the variation in sound pres-unwanted particles from an object by generating

sure with the working distance was measured and thehigh-frequency mechanical oscillations in water or a sol-

optimal working distance was determined for the clean-vent. The cleaning mechanism is based on cavitation

ing test. Figure 2 shows images of the UWF device.

and vibrational acceleration within the washing fluid.

Since the UWF device is to be used in the mouth for

Ultrasonic cleaners that operate at frequencies below

this application, the water flow rate was chosen to be

100 kHz primarily depend on this cavitation mecha-

approximately 300 ml/min. Although it was observed

nism12）. This type of ultrasonic cleaner is suitable forthat the water flow remained rectilinear, it developed

removing particles with sizes in the order of several

into a spray after a working distance of ≥ 60 mm

microns and is used, for example, for cleaning optics（Fig. 2）.

and precision parts. Cleaners in the MHz range are

2 ．Sound pressure simulation

operated at very high frequencies above 1 MHz, andFigure 3 shows a simulated ultrasonic beam patternthese depend on the vibrational acceleration mecha-

under the free-boundary condition in water. The ultra-

nism 13）. At these frequencies, particles measuringsonic field was calculated using the ring function≤ 0.3μm in diameter can be removed . This type of

method18）at a frequency of 400 kHz（wavelength 3.75

cleaner relies on high-frequency acceleration force andmm）, a transducer size of φ＝30 mm, and a soundis used, for example, for cleaning photo masks or siliconvelocity of 1,500 m/s. The maximum sound pressure

14‒16）

area appears around 50‒80 mm from the transducer.

14）

wafers in the manufacture of semiconductors.

Dec, 2021

Utility of 400 kHz Ultrasonic Water Flow to Removal of Biofilm
ultrasonic transducer

61

This simulation shows the profile of the sound pressurethat we experimentally measured using hydrophones.

ultrasonic wave

washing machine bodywater

3 ．Measurement of sound pressure
The sound pressure distribution in the UWF system
was measured using a hydrophone（ONDA HNR - 0500;
with ‒250 kHz to 1 MHz calibration）. Figure 4 shows
the waveform of the sound pressure p0－p of the UWF

washing fluid

system.

specimen

The measurements were performed for various input
settings and various distances from the transducer.

Fig. 1In the present experiment, the sound pressure was

The UWF systemmeasured as a function of working distance for various
input powers, and an optimal condition was determined
for the cleaning test.
4 ．Cleaning experiment for artificial plaqueThe cleaning effects of various input powers were

Fig. 2 Images of UWF
1：Water flow, 2：Ultrasonic water flow, 3：Ultrasonic water flow（Close-up view）

0.40MHz（WaveLength：3.75mm）

20
10
0

X（mm）

d：30.00 mm

－10
－20
0

50

100
Z
（mm）

150

200

2.00Pp（max2.00Pp）

Fig. 3 Simulated beam pattern in the free-boundary condition
1：Ultrasonic transducer, 2：Maximum sound pressure of the ultrasonic wave, 3：Level of soundpressure. The maximum（peak）pressure（Pp）is 2.00.

ODEP Vol. 1, No. 162p0－p：0 to（positive）peak pressurep0－p：0 to（negative）
peak pressure

Fig. 4

Waveform of the sound pressurebiofilm formation and removal using UWF treatment.
The study protocol was approved by the ethics committee of the Showa University School of Dentistry, andwritten informed consent was obtained from all participants（No. 2011‒013）.
The surface characteristics of the titanium specimens
were similar to those of commercial dental implants（Setio, GC Corporation, Japan）. Two types of surface
Fig. 5 Titanium test plate placed in the mouth
Plaque biofilm was formed on the titanium
specimens in situ over a period of 72 h（roughsurface and mirror-finished surface, 5 mm×7

were examined, a mirror-finished surface and a sand-mm）at the buccal site of the molar and premolar teeth on custom-made stents.

2 ）Estimation of the area covered by the residual

blasted rough surface. The sandblasted surface washydrophilic, grade IV, with a surface roughness Ra＝2.54 μm and Rz＝11.87μm.
plaque biofilm
The titanium specimens were subjected to water

investigated using artificial plaque on a titanium speci-

flow alone or to UWF, and in each case the area cov-

men. The input power to the ultrasonic transducer

ered by residual plaque biofilm（RPB）was measured as

ranged from 5 to 25 W, and the washing time was 60 s.

the percentage of the area of a randomly selected

The area of artificial plaque remaining on the titanium

scanned surface using a digital microscope（VHX - 2000,

specimen after cleaning was measured using a digital

KEYENCE, Japan）. To compare the RPBs, the initial

optical microscope. The residual artificial plaque（RAP）

area covered by the plaque biofilm was defined asin the scanned area was defined as the percentage of

100％ RPB20）.

the area that retained the artificial plaque after clean-

6 ．Scanning electron microscope obser vations

ing. This area was measured by setting a threshold

After the application of UWF, the titanium speci-

value for the color.

me ns w e re f ixe d in g lu t ar a ld eh y d e［2 .5％ phos-

5 ．Cleaning experiment for biofilm

phate-buffered saline（PBS）］for 30 min and then rinsed

1 ）Biofilm formation on titanium specimens

in PBS. Then, the samples were fixed in osmic acid（1％Figure 5 illustrates the position of the titanium plates

PBS）for 30 min at 4℃ and subsequently dehydrated in

fixed to a removable custom-made stent placed in thea graded ethanol series（50％‒100％ for 10 min）. Finally,

mouth of a volunteer. The specimens were placed at

the samples were dried in a critical point drying

the buccal sites of molar and premolar teeth in themachine（HCP-2, Hitachi, Japan）. All samples were

mouths of five male subjects for 72 h to allow plaquemounted on scanning electron microscope（SEM）sam-

19）

and were removed only

ple stubs（E-1030, Hitachi）and sputtered with platinum.

during eating or brushing. They were then analyzed for

The plaque biofilm was analyzed using an S - 4700 SEM

biofilm to form on themDec, 2021

Utility of 400 kHz Ultrasonic Water Flow to Removal of Biofilm
1.00

Sample size was calculated using EZR（Saitama Medical Center, Jichi Medical University, Japan）and by
performing power analysis for RPB on titanium specimens. The sample size was calculated for a＝0.05 andb/a ratio＝0.8, and the effect of size was calculatedusing the mean and SD of the variables. The analysis
indicated that a sample of five subjects was needed.

Sound pressure［MPa］（Hitachi）at×350 magnification.
7 ．Sample size calculation

0.80
0.60
0.40
0.20
0.00
－0.20
－0.40
－0.60
－0.80

8 ．Statistical analysis－1.00

The results are presented as median values with the10

top 25th and the bottom 25th percentiles（first andwhich is a graphical user interface for R（The R Foun-

20

30

40

50

60

Distance［mm］

third quartiles）rather than as mean and standard deviation. Statistical analyses were performed using EZR,

63

positive peakFig. 6A

dation for Statistical Computing, Vienna, Austria）. The
Mann-Whitney U test was used to evaluate the differ-

negative peak

Axial position and sound pressure fieldmeasured in the water flow using ahydrophone

ences in RPB, and the Wilcoxon signed-rank test was
1.50

biofilm（IPB）and RPB. The correlation was analyzed
using Spearman’s correlation test. A p value of ＜0.05
was considered to be significant.

Results
Figure 6 depicts the relationship between the negative and positive peak pressures of the sound pressure
waveform in the UWF system. The sound pressure
measurements were confirmed to be reasonably accu-

Sound pressure［MPa］

used to compare differences between the initial plaque

1.00
0.50
0.00
－0.50
－1.00
－1.50

rate based on the calculated results of the beam pat-ultrasonic transducer（Fig. 6A）. Therefore, the work
distance for the artificial plaque cleaning test was cho-

10

15

20

25

Input power［W］

tern distribution（Fig. 6A）. An increase in sound pressure can be observed at distances ＞50 mm from the5positive peakFig. 6B

negative peakSound pressure-input curves obtained byhydrophone

sen as 50 mm. The absolute values of the negative andpositive peak pressures were approximately equal. As

mm, the area cleaned by the UWF was considerably

shown in Figure 6B, when the input setting waslarger. Figure 7B shows the percentages of RAPs forchanged from 5 to 25 W, the sound pressure increased

various input powers. Water flow without ultrasonic

depending on the input power. For example, the maxi-

waves was unable to remove any of the artificial

mum sound pressure was 0.6 MPa when the input

plaque. As shown in Figure 7B, when the input setting

power was 15 W.

was changed from 5 to 25 W, the RAP declined with an

Figure 7A shows a painted titanium specimen before

increase in the input power; for example, the RAP was（left）and after（right）cleaning. The input power to the

60％ when the input power was 15 W. There was aultrasonic transducer was ‒5‒25 W, and the cleaning

negative correlation between the RAP rate and thetime was 60 s. As depicted in Figure 7A, metallic colors

input power. Based on these results, the device condi-

indicate the areas from which artificial plaque had been

tions used for evaluating the removal of dental biofilm

removed. Although the nozzle outlet diameter was 6

were set as follows: the input power was 15 W and the

ODEP Vol. 1, No. 1

64
Beforedistance from the transducer to the specimen surface

After

was 50 mm.
Figures 8A and 8B show optical images of the initial

0W

titanium surface. Both types of specimen（i.e., those witha mirror-finished surface and those with a rough surface）exhibited an erythrosine-stained plaque biofilm
coverage of 100％ after 72 hours of intraoral exposure（Figs. 8C and 8D）. The titanium specimens were sub-10W

jected to UWF under the abovementioned conditions,and digital microscopic photographs of each surface
revealed the removal of plaque biofilm（Figs. 8E and8F）. Biofilm removal was observed after approximately
45 s from the initiation of UWF. Plaque removal was
found to increase in a time-dependent manner for up to

15W

180 s. All stained specimens were improved similar tothat in the initial titanium surface.
The total RPBs after cleaning on the mirror-finishedand rough surfaces were 22.8％（median）and 7.05％（median）, respectively（Fig. 9）. A significant decrease

20W

in RPB after the application of UWF was confirmed bycomparison with the surfaces before treatment. Eightspecimens were included in each group.
The clean titanium specimens showed a flat（Fig.
10A）or sharp-edged surface（Fig. 10B）. The specimens

25W

exhibited a thick dental biofilm（Figs. 10C and 10D）; in
particular, the specimen with a rough surface wasentirely coated（Fig. 10D）. The plaque biofilm could not

Fig. 7A Cleaning experiment for artificial plaquebe removed by normal water flow treatment, and it

100
80
RAP（％）

remained intact with only a slight disturbance of the

r＝－0.955

surface（Figs. 10E and 10F）. However, the UWF treatment removed not only the plaque biofilm but also the60

water-insoluble glucans from both types of surface（Figs. 10G and 10H）. The grooves and peaks on the40

rough titanium surface were visible after the UWF
treatment（Fig. 10H）. There were no alterations in the20
0

surfaces of the specimens treated by UWF.
0

5

10

15

20

25

Discussion

Input power（W）
Correlation of the RAP areas and input
power
The cleaning effects of various input powers
were investigated using artificial plaque on a titanium specimen. The input power to the ultrasonictransducer ranged from 5 to 25 W, and the washing time was 60 s. RAP declined with increase in
the input power. There was a negative correlation
between the RAP and the input power.
Fig. 7B

Sound pressure estimation based on input power
Regarding the estimation of sound pressure from the
input power, sound pressure, p0－p , is represented using

the electric input power E（W）and the electromechanical coupling factor, η（％）, as follows:
p0－p＝1.4×

E×η2×ρc ……（1）S
where, S is the effective area of the transducer（m2）,Dec, 2021

Utility of 400 kHz Ultrasonic Water Flow to Removal of Biofilm

65

Fig. 8 Optical images of plaque biofilm removal by the ultrasonic water flow
Control specimens of titanium with a mirror-finished surface（A）and a rough surface（B）.
Plaque biofilm formed on the titanium surfaces after 72 h（C, D）. Surfaces after treatment by
ultrasonic water flow for 3 min（E, F）. Magnification：×100＊＊

pressure 0.23 MPa becomes 0.59 MPa at the focal point.

100

The measured value was 0.6 MPa.

RPB
（％）

80

Therefore, the acceleration α associated with a fre-60

quency of 400 kHz is determined using equation（3）asfollows:40

α＝－2πf20
0

From Figure 4, if we assume that the sound pressure
is around 2.5 times greater at the focal point, the soundpre treatment mirror surface rough surface

The RPB areas after ultrasonic water
treatment of the specimens
The total RPBs are 22.8％（median）and 7.05％（median）
on the mirror-finished and rough surfaces,
respectively. There is a significant decrease in the
RPB when compared to its value before treatment
for both the mirror-finished and rough surfaces.
＊: p＜0.05, Mann-Whitney U test（n＝8）.
Fig. 9p0－p＝－2π×400×103×ρc
6
2）……（3）－1.0×10（m/s

0.6×106
1.5×106＝

This value is approximately 100,000 times the gravitational acceleration g（＝9.8 m/s2）.
The mechanism involved in ultrasonic cleaning in the
MHz range is related to the magnitude of vibrational
acceleration in the washing fluid13）. We combined anultrasonic cleaner at 400 kHz with flowing water toproduce an UWF system that is capable of effectively
removing dental biofilm from titanium surfaces. The
enhanced cleaning effect of this setup may be due toρ＝1×103kg/m3 is the density of water, and c＝1,500

the sound pressure derived from vibrational accelera-

m/s is the speed of sound in water. If the effectivetion in the fluid.

diameter of the transducer is assumed to be 0.02 m andIn the simulation illustrated in Figure 3, a region ofelectromechanical coupling factor, η, is assumed to behigh sound pressure is observed at distances of ≥50

0.5, at the electric input power E＝15 W, we obtain

mm from the transducer, and the beam pattern of thep0－p＝1.4×

3.75－6

201×10× 1.5 × 106=0.236（MPa）……（2）high-frequency ultrasound has a narrow directivity
angle, which is characteristic of high-frequency ultrasonic cleaners. In addition, the sound pressure measurements depicted in Figure 6 indicate that the soundODEP Vol. 1, No. 1

66

Fig. 10 SEM micrographs of plaque biofilm on titanium surfaces
Control specimens of titanium with a mirror-finished surface（A）and a rough surface（B）. Untreated titanium
surfaces covered by dense bacterial biofilm（C, D）. Micro-structured titanium surfaces treated with a water jet
demonstrating little or no disruption of the bacterial chain（E, F）. Titanium surfaces after exposure to ultrasonicwater flow for 3 min, resulting in complete disruption of the bacterial chain and also removal of the water-insoluble glucans（G, H）（Magnification：×350）.

pressure reaches its maximum value at a distance of 50

due to the vibrational acceleration of the washing fluid.

mm from the transducer, and when the input power is

It is thought that the vibrational acceleration and wash-

set to 15 W, the sound pressure is also stable. This sta-

ing fluid exert a synergistic effect on the biofilm

ble sound pressure could be measured in all directions

removal. The result of smaller RPB areas on UWF-

if sufficient flowing water was obtained. Therefore, thistreated rough surfaces may have been caused by ultra-suggests that the same effect can be expected in thesonic waves propagating in the thread gap. In addition,

human oral cavity. However, it is considered that a noz-the ultrasonic waves may have reflected off the unevenzle tip needs to be developed to transmit the ultrasonicsurface and effectively peeled off the biofilm.

energy of the flowing water to a narrow part of theSEM observations of both mirror-finished and roughoral cavity. Hence, these values were determined to be

surfaces showed that titanium surfaces covered by den-

the optimal conditions for the experiment.

tal biofilm recovered almost to their original state in

The efficiency of ultrasonic cleaning depends on theterms of both biofilm content and surface characteris-

sizes of the particles to be removed and the frequency

tics after the UWF treatment. The results also demon-of the ultrasound. Low-frequency（‒28‒200 kHz）ultra-

strated that even the biofilm from the crests andsonic cleaners are suitable for removing particles withtroughs of threaded implants could be removed using≥10μm diameter, and high-frequency（‒1 MHz）ultra-

the proposed cleaning system（Fig. 10）, whereas con-sonic cleaners can easily remove much smaller particles

ventional devices such as hand scalers, toothbrushes,measuring ≤0.3μm in diameter14,21,22）. The 400 - kHz

and ultrasonic scalers cannot reach such minute struc-ultrasonic cleaner used in this study is commonlytures since the diameters of the working parts are gen-applied to remove particles measuring ‒0.2‒5μm in

erally too large.

23）

diameter, corresponding to the size of bacteria . Fur-

In general, the damage caused by ultrasonic cleaners

thermore, oral bacteria are electrostatically attached todepends on their frequency, with higher frequencies

the tooth or implant surfaces and can be removed bycausing greater damage. In this study, there were no

vibrational acceleration. Therefore, we suggest that achanges to the titanium surfaces treated using thehigh-frequency or a midrange-frequency ultrasonic

UWF system.

cleaner is most suitable for removing plaque biofilm.

Cleaning procedures with conventional devices,

The use of UWF was effective in removing biofilm

which require contact with the dental implant, canfrom titanium surfaces（Fig. 9）. This removal effect wasincrease the surface roughness of titanium, therebyDec, 2021

Utility of 400 kHz Ultrasonic Water Flow to Removal of Biofilm

affecting microbial colonization and inducing the forma-

67

bacterial patterns present at implant and tooth sites in. Our proposed technique avoids thissubjects on supportive periodontal therapy. Clin Oral

Therefore, the advantages of the UWF system can4）Rams TE, Link CC Jr. Microbiology of failing dental
implants in humans: electron microscopic observations.
J Oral Implantol 1983; 11: 93‒100.
5）Kalykakis GK, Mojon P, Nisengard R, Spiekermann H,
Zafiropoulos GG. Clinical and microbial findings on
osseo-integrated implants; comparisons between partially dentate and edentulous subjects. Eur J Prosthodont Restor Dent 1998; 6: 155‒159.
6）Lindhe J, Meyle J, Group DoEWoP. Peri-implant diseases: Consensus Report of the Sixth European Workshop on Periodontology. J Clin Periodontol 2008; 35:282‒285.
7）Heitz-Mayfield LJ. Peri-implant diseases: diagnosis andrisk indicators. J Clin Periodontol 2008; 35: 292‒304.
8）Albouy JP, Abrahamsson I, Persson LG, Berglundh T.
Implant surface characteristics influence the outcome oftreatment of peri-implantitis: an experimental study indogs. J Clin Periodontol 2011; 38: 58‒64.
9）Claffey N, Clarke E, Polyzois I, Renvert S. Surgical treatment of peri-implantitis. J Clin Periodontol 2008; 35:316‒332.
10）Sato S, Kishida M, Ito K. The comparative effect of ultrasonic scalers on titanium surfaces: an in vitro study. J
Periodontol 2004; 75: 1269‒1273.
11）Baek SH, Shon WJ, Bae KS, Kum KY, Lee WC, Park YS.
Evaluation of the safety and efficiency of novel metallic
ultrasonic scaler tip on titanium surfaces. Clin Oral
Implants Res 2012; 23: 1269‒1274.
12）Morita K. Ultrasonic cleaning. Kindai Henshu, Tokyo
1989.
13）Kim JO, Choi S, Ho Kim J. Vibroacoustic characteristics
of ultrasonic cleaners. Applied Acoustics 1999; 58: 211‒
228.
14）Mayer A, Shwartzman S. Megasonic cleaning: A newcleaning and drying system for use in semiconductor
processing. J Electron Mate 1979; 8: 855‒864.
15）Busnaina AA, Gale GW. Ultrasonic and megasonic particle removal. Proc Precis Clean 1995; 15: 347‒359.
16）Hatano H, Kanai S. High-frequency ultrasonic cleaningtank utilizing oblique incidence. IEEE Trans Ultrason
Ferroelectr Freq Control 1996; 43: 531‒535.
17）Honda Keisuke. Ultrasonic cleaning device. U. S Patent
4,834, 124 May 30, 1989.
18）Ohtsuki S. Calculation method for the nearfield of asound source with ring function. J Acoust Soc Jpn 1974;
30: 76‒81.
19）Rimondini L, Fare S, Brambilla E, Felloni A, Consonni C,
Brossa F, Carrassi A. The effect of surface roughness
on early in vivo plaque colonization on titanium. J Peri-

10,11,24）

tion of plaquedisadvantage.

be summarized as follows: 1）it can remove dental biofilm from minute structures, 2）the use of a higher frequency in this system implies that there is less cavitation damage, and 3）it is a non-contact cleaning systemthat does not cause additional damage. However, afuture task is to reduce the size of the UWF device andthe amount of water as the ultrasonic transducer islarge.

Conclusion
We demonstrated a novel UWF cleaning mechanismthat can efficiently remove plaque biofilm without altering the characteristics of the titanium surface. In our
experiment, titanium surfaces were exposed to UWF
with a water flow rate of 300 ml/min, an ultrasonic frequency of 400 kHz, and an input power of 15 W（soundpressure of 0.6 MPa）
. The proposed UWF mechanism
can be applied to devices for the maintenance of dental
implants.
Acknowledgments
This work was supported in part by a Grant-in-Aid for
Exploratory Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan（24659927 to.
T. T.）
The authors would like to thank Enago（www.enago.jp）
for the English language review.

Conflict of Interest
The authors declare they have no financial interest, either
directly or indirectly, in the products or information
described in this article.

References
1）Costerton JW, Montanaro L, Arciola CR. Biofilm in
implant infections: its production and regulation. Int J
Artif Organs 2005; 28: 1062‒1068.
2）Costerton JW, Montanaro L, Arciola CR. Bacterial communications in implant infections: a target for an intelligence war. Int J Artif Organs 2007; 30: 757‒763.
3）Agerbaek MR, Lang NP, Persson GR. Comparisons of

Implants Res 2006; 17: 18‒24.

68
odontol 1997; 68: 556‒562.

ODEP Vol. 1, No. 1
22）Kashkoush I, Busnaina A. Submicron particle removal

20）Schwarz F, Sculean A, Romanos G, Herten M, Horn N,
Scherbaum W, Becker J. Influence of different treat-using ultrasonic cleaning. Particulate Science and Technology 1993; 11: 11‒24.

ment approaches on the removal of early plaque biofilms and the viability of SAOS2 osteoblasts grown ontitanium implants. Clin Oral Investig 2005; 9: 111‒117.
21）Busnaina A, Taylor J, Kashkoush I. Measurement of theadhesion and removal forces of submicrometer particles
on silicon substrates. J Adhes Sci Technol 1993; 7: 441‒
455.

23）Listgarten M. Structure of the microbial flora associatedwith periodontal health and disease in man: A light andelectron microscopic study. J Periodontol 1976; 47: 1‒18.
24）Shafagh I. Plaque accumulation on cast gold complete
crowns polished by a conventional and an experimental
method. J Prosthet Dent 1986; 55: 339‒342.

Dec, 2021

69

Original Article

ODEP 1（1）：69～76，2021

Effect of Root Canal Transportation by Minimally Invasive Endodontic
Shaping of Canal Orifice Dentin
Kazuyuki TSUBAKI, Mai UTSUNOMIYA, Kaori SHIMOJIMA,
Noriko MUTOH and Nobuyuki TANI-ISHII
Department of Pulp Biology and Endodontics, Kanagawa Dental University

Abstract
Background: The purpose of this study was to analyze the effect of minimally invasive（MI）shaping of
canal orifice dentin by evaluating the shaping characteristics of TruNatomy（TN）, which was developed as a
Ni-Ti file capable of MI endodontics.
Methods: A J-shaped canal model was used to compare prepared final canal morphology using a minimallyinvasive shaping TN file and a conventional shaping ProTaper NEXT（PTN）file. The TN files used in theexperiment were classified into two groups: no straight-line access（group A）and straight-line access（group
B）. The PTN files of the control were also classified into two groups: no straight-line access（group C）andstraight-line access（group D）, and the increase in canal width and the median displacement（transportation）
were compared.
Results: Analysis of the canal width by TN showed that the displacement on the inner side increased by 5
mm in both groups A and B, but the displacement on the outer side increased at the other measurementsites. In groups C and D using PTN, the displacement on the inner side increased by 3‒5 mm. On the other
hand, the displacement on the outer side increased by 8 mm, and the displacement in group D increased. The
canal transportation using TN was 0.1 mm or less in both groups A and B, although preparation by PTN
showed significant transport to the inner side by 3‒5 mm and outer side of 8 mm in group D compared togroups A and B.
Conclusion: TN shaping was shown to preserve tooth structure and canal geometry without straight-line
access. It was confirmed that TN, which was developed based on the concept of MI Endo, enables accurate
root canal shaping and reduces transportation.
Key words: canal transportation, shaping ability, straight-line access, TruNatomy, ProTaper NEXT

Corresponding author: Nobuyuki TANI-ISHII, Department of Pulp Biology and Endodontics, Kanagawa Dental University, 82,
Inaoka-cho, Yokosuka, Kanagawa 238‒8580, Japan
TEL: ＋81‒46‒822‒9527, FAX: ＋81‒46‒822‒8856, E-mail: n.ishii@kdu.ac.jp
Received for Publication: September 7, 2021/Accepted for Publication: October 12, 2021
DOI: 10.11471/odep.2021‒010

ODEP Vol. 1, No. 1

70

be a prerequisite for improving the prognosis of endodontic treatment because it enables apical closure by

Introduction

proper root canal filling by maintaining the apical root
It has been reported that endodontic treated teeth

canal morphology during root canal preparation23）.

have a higher risk of fracture depending on the mor-

However, next-generation Ni-Ti files, microscopy, andphology and amount of residual tooth substance1‒3）. Seo

cone-beam computed tomography enable dentists to4）reported that 79％ of vertical root canal frac-make access cavities as minimally invasive as possible

tured teeth were endodontic treated teeth. Endodontic

and maintain the dentin structure of the tooth being

treated teeth and root canal fractures have been shown

treated.

et al.

to be related to stress during root canal shaping5）. Den-

The TruNatomy（TN; Dentsply Sirona, Ballaigues,

tin cracks that occur during root canal shaping are

Switzerland）file was designed based on the concept ofthought to be the cause of vertical root canal fracture

MI Endo and was developed to enable accurate root

along with long-term disinfectant administration in thecanal shaping without straight-line access, and rotatesroot canal and excessive vertical filling pressure during

at high speed. It is a unique thin Ni-Ti wire with a max-

root canal obturation6）.

imum flute diameter of 0.8 mm instead of 1.2 mm,

In recent years, minimally invasive endodontics（MIwhich is commonly used in other Ni-Ti files. Also, it has

Endo）has been advocated, and root canal shaping bya regression taper and an off-centered parallelogramminimal dentin removal that preserves the dentin on

cross section in the file system as used by Peters etthe canal orifice reduces root canal fracture7, 8）. In theal.24）. The TN file consists of a file system designed

basic process of canal shaping, straight-line access isaccording to the conventional ProTaper NEXT（PTN,

one of the most important procedures and is a prereq-

Dentsply Sirona）file. Since the PTN file25）is used based

uisite step for shaping and obturation9‒11）. The removalon the conventional root canal shaping concept thatof infected dentin at the canal orifice by straight-line

requires straight-line access, it can be compared withaccess and reduction of the degree of root canal curva-

the minimally invasive shaping TN file as a control forture facilitate accurate root canal shaping. However,this study. The physical characteristics of PTN files

straight-line access has been reported to be associatedhave been reported to be extremely flexible26, 27）. Simi-

with excessive shaping of the orifice dentin, reducing

lar to PTN, the TN file is constructed with a cross-sec-

both dentin stiffness and fracture resistance12, 13）. It has

tional form in which the file cutting surface is off-cen-been reported that preservation of orifice dentin of thetered, but the characteristics of the alloy are different.

upper one-third of the root canal is more important

Although the tapers of both TN and PTN files are vari-than preservation of crown occlusal surface dentin toable, both have a 6％ taper for the apical 3 mm. In addi-

prevent root canal fracture 14, 15）. It has also beention, the file system has a similar format, but the PTN

reported that contouring of the access cavity by MI

file tip sizes are X1（#17）, X2（#25）, and X3（#30）,

Endo is related to the structural maintenance of thewhile the TN file tip sizes are Small（#20）, Prime（#26）,

crown dentin, but the relationship between minimallyand Medium（#36）. Both file systems are usuallyinvasive access to the cervical dentin and the fracture

designed to complete root canal shaping with two files:

resistance of the root is controversial16‒18）. However,X1 and X2 for PTN, Small for TN, and Prime for cases

root canal shaping by MI Endo of orifice dentin isof pulpectomy in the root canal.

reported to maintain high fracture resistance compared19）

to root canal shaping by straight-line access .

The purpose of this study was to evaluate the characteristics of canal preparation by TN based on the MI

Currently, MI Endo20） is being used mainly on the

Endo concept. For the evaluation, root canals were pre-west coast of the United States in order to prevent rootpared using a minimally invasive cutting type TN and acanal fracture, unlike the conventional root canal prepa-

conventional cutting type PTN, and the total amount ofration method. MI Endo aims to maximally preserve

removed root canal wall and the canal transportation

dentin and also influences caries diagnosis, treatment

were measured and compared. The results clarified the

choices, access cavity design, and root canal or resto-

relationship between straight-line access during root

ration choices

21, 22）. The access cavity is considered tocanal preparation and final canal morphology.

Dec, 2021

Canal Transportation by Straight Line Access

Materials and Methods
1 ．Operator and test canal model
The canal preparation was performed by a single
operator（specialist of JEA and JCD）using Ni-Ti（PTN）
files in strict accordance with the manufacturer’s rec-

71
D16
D12
D6
D0
0.79
0.73
0.53
0.20

A）

0.80

0.73

0.49

0.26

C）

1.16

0.91

0.49

0.17

D）

1.20

1.04

0.63

0.25

B）

ommendations for each system.
For the experiment, a total of 40 J-type root canalmodels（curvature 30°, apex diameter #15, taper 02,
canal length 16.5 mm: End Training block canals,
Dentsply Sirona）made by epoxy resin were used. All
root canal models were attached to the Endo Training
Model Castillo（VDW, Frankfurt, Germany）, and root

Fig. 1The dimensions of TN（A：Small 20/04 V，B：
Prime 26/04 V）and PTN（C：X1 17/04，D：
X2 25/06）files

canal preparation was performed in a situation where
the root canal morphology could not be confirmed.

path was formed by TN Glider. Root canal preparation

2 ．Root canal preparation

by TN was shaped using X Smart plus（Dentsply

To analyze the effect of straight-line access on theSirona）by the following procedure. For root canal

final canal morphology after root canal preparation, TN

preparation, shaping was performed with TN Smalland PTN files were compared. The TN files used in the（20/04V）, then TN Prime（26/04V）, with both files set

experiment were classified into two groups: no straight-

to 500 rpm and 1.5 Ncm, and three vertical movement

line access（group A）and straight-line access（group

operations were performed. The working length was

B）. The PTN files of the control were also classified

reached in 3 cycles as 1 cycle, then root canal prepara-

into two groups: no straight-line access（group C）andtion was completed by cleaning the root canal with 2 ml

straight-line access（group D）, and the increase in rootof distilled water（DW）at the end of each cycle.

canal width and the canal transportation were com-

In the control group using PTN, penetration to thepared. In each experimental group, the PTN Accessapex with K file #10 was confirmed before the start ofOpener XA of the PTN file system was used for theroot canal preparation. After confirmation of penetra-

straight-line access group, and the Orifice 5（OM）of thetion, the group was classified into no straight-line group

TN file system was used for the no straight-line group

C using Orifice Modifier 20/08, and straight-line groupto enlarge the root canal orifice.

D using PTN Access Opener XA. Then, a glide path

In each experimental group, the final preparation

was formed in the TN Glider group（n＝20）. Root canalmorphology was set to TN Prime（file tip diameter

formation by PTN was performed using X Smart plus

#26/average taper 04（Variable: V）: ISO #26/04V）andby the following procedure. For root canal preparation

PTN file X2（ISO #25/06）. Figure 1 shows the dimen-

by PTN, two X1（17/04）and X2（25/06）files weresions of the TN and PTN files, and Table 1 shows theused, with both files set to 300 rpm and 2.0 Ncm. The

files used up to the root canal preparation in eachfirst X1 and second X2 files were penetrated to theexperimental group.

working length three times, then root canal preparation

For root canal preparation by TN in the experimental group, after confirming penetration to the apex witha stainless-steel K file #10（Dentsply Sirona）before thewas completed by cleaning the root canal with 2 ml of
DW at the end of each cycle.
3 ．Evaluation of root canal morphology

start of root canal preparation, straight-line access was

The amount of root canal morphological displace-made using Orifice Modifier 20/08 to shape the canalment was analyzed by measuring the total amount oforifice. The experimental TN group was classified into

removed root canal wall on the outer and inner sides of

no straight-line group A using Orifice Modifier 20/08

the curved root canal. A stereoscopic microscope

for the root canal orifice opening, and straight-line（SZX16; Olympus, Tokyo, Japan）and a digital cameragroup B using TN Access Opener XA. Then, a glide（DP71; Olympus）were used for measurement, transpar-

ODEP Vol. 1, No. 1

72
Table 1

The files used up to root canal preparation in each experimental group

Experimental

Ni‒Ti file system

Group

Straight‒line（＋/－）

Scout

Orifice open

Glide path

Preparation

Preparation

Start file

Final fileA

TruNatomy
Straight‒line（－）#10

O. Modifier 20/08

Glider 17/02 V

Small 20/04 V

Prime 26/04 V

B

TruNatomy
Straight‒line（＋）#10

A. Opener XA

Glider 17/02 V

Small 20/04 V

Prime 26/04 VC

ProTaper NEXT
Straight‒line（－）#10

O. Modifier 20/08

Glider 17/02 V

X1 17/04

X2 25/06

D

ProTaper NEXT
Straight‒line（＋）#10

A. Opener XA

Glider 17/02 V

X1 17/04

X2 25/06

A）

B）

C）

D）

＊
8mm

＊
＊＊

＊＊

5mm
3mm
2mm
1mm
Apex

Fig. 2 Superimposition of pre‒ and post‒preparation images
TN no straight‒line group（A），TN straight‒line group（B），PTN no straight‒
line group（C），and PTN straight‒line group（D）．＊：Straight‒line access，＊＊：Final preparation by PTN X2 file.

ent root canal models pre- and post-preparation were

sis between two groups）.

superimposed on digital images, and the obtained image
data was captured on a PC for measurement. Measurements were performed using software（WinRoof; Tokyo,

Results

Japan）（Fig. 2）. To measure the amount of root canal

The root canal morphology in post-preparation waswall shaping in the root canal model, the positions were

evaluated by measuring the increase in root canalset at 1, 2, 3, 5, and 8 mm from the apex as the measur-width and the median displacement on the inner anding unit, and the root canal width was increased onouter sides at positions 1, 2, 3, 5, and 8 mm from theeach of the outer and inner canal sides. The distanceapex. Comparing the prepared canal morphology（Fig.

from the root canal wall of pre- to post-preparation and2）of the canal models in pre- and post-root preparation

the median root canal value as canal transportationof groups A, B, C, and D, flare formation on the canalwere measured and statistically processed.

orifice was not observed in group A without straight-

4 ．Statistical processing

line access. However, in group C in which straight-line

The measured values of root canal wall cutting wereaccess was not formed, flare formation on the canal ori-

statistically processed with a risk factor of 5％ using

fice was observed similar to that in groups B and D in

one-way ANOVA, Bonferroni-Dunn multiple compari-

which straight-line access was formed. Furthermore, in

son test, and Mann-Whitney U test（comparative analy-

group D with straight-line access, an increasing ten-Dec, 2021

Canal Transportation by Straight Line AccessMeasurement point（mm）

8
＊
＊
＊
＊

5

＊
＊

3

2

0.4

0

0.1

Total amount of removed resin（Mean±SDmm）

0.5

0.4

0.3

0.2

Outer side
0.2

0.3

1
0.1

0.4

＊

0.2
0
－0.2
－0.4

＊

－0.6
1

2

3

5

8

Measurement point（mm）

0

0.5

＊

0.6

－0.8

1
Inner side
0.6

PTN（＋）（A） TN（－）
（B）
,
TN（＋）
（C）
,
PTN（－）（D）,
0.8

Canal transportation（mm）（A） TN（－）
（B）
,
TN（＋）
（C）
,
PTN
（－）
（D）
,
PTN（＋）

73

Total amount of removed canal wall（mm）atthe different levels in post‒preparation
TN no straight‒line group（A），TN straight‒line
group（B），PTN no straight‒line group（C），and PTN
straight‒line group（D）．＊：Significant difference（p＜0.05）.
Fig. 3

Canal transportation at different levels from
apex in post‒preparation
TN no straight‒line group（A），TN straight‒line
group（B），PTN no straight‒line group（C），and PTN
straight‒line group（D）．＊：Significant difference（p＜0.05）between TN（A，B）and PTN（D）.
Fig. 4

Discussion

dency was observed in the amount of cutting on the

Regarding endodontic treated teeth, it has been sug-

inner side at the start position of the curve of 3‒5 mm.

gested that the treatment process itself such as access

The results of measuring the total amount ofcavity preparation and root canal enlargement has aremoved root canal wall are shown in Fig. 3 and canalrisk of causing root canal fracture, and in particular, atransportation is shown in Fig. 4 for all experimental

relationship between the distribution of occlusal stressgroups.

and root canal fracture has been reported28）. To reduce

According to the results of analyzing the increase in

the risk of root canal fracture, the concept of MI Endo

total amount of root canal wall removed by TN, the dis-with minimal access cavity and root canal preparation

placement on the inner side increased at the measure-using Ni-Ti files is recommended. It is essential to nego-ment site by 5 mm at the start position of root canaltiate the root canal using a K file at the start of root

curvature in both groups A and B, but the displace-canal preparation, to obtain anatomical root canal infor-

ment on the outer side increased at other measurement

mation such as curvature, calcification, apical foramensites. On the other hand, in both groups C and D, thediameter, and root canal length. Furthermore, whendisplacement on the inner side at the measurement site

preparing curved root canals, removal of the orificeincreased by 3‒5 mm, on the outer side by 8 mm, anddentin ridge is considered to be the most importantthe displacement of group D increased in particular

step, facilitating file insertion to the apical foramen.

（Fig. 3）.

That is, the removal of dentin ridges by straight-line

The median displacement as canal transportation byaccess and the establishment of glide paths have been

TN was 0.1 mm or less at all measurement sites in bothregarded as essential techniques for the safe use ofgroups A and B. The median displacement by PTN was

Ni-Ti files.

significant: 3‒5 mm to the inner side at the measure-On the other hand, it has been reported that straight-

ment site in group D compared to groups A and B, andline access, which has been considered the basis of root

8 mm to the outer side. On the other hand, the mediancanal preparation, reduces fracture resistance anddisplacement of group C was not significantly different

causes root canal fracture due to excessive cutting offrom that of groups A and B（Fig. 4）.

orifice dentin. Endodontic treated teeth have highODEP Vol. 1, No. 1

74stress concentration in the cervical region 29, 30）, andaccess group（D）revealed interesting results. The

excessive cutting of orifice dentin is considered to be aincrease in root canal width and the median displace-factor that increases the susceptibility to fracture .

ment in the group without PTN straight-line access（C）

These study reports have shown that minimizing flare

showed the same amount of displacement of 1‒3 mm atformation due to straight-line access may reduce thethe apex as in groups A and B by TN. Furthermore,risk of cervical fracture. In addition, a straight-line

the displacement by TN in groups A and B increased

31）

access unformed root canal causes stress dispersion in

by 5 mm only on the inner side and by 8 mm on thethe orifice dentin when normal stress is applied to theouter side, but no significant difference was observed,

occlusal surface, whereas the straight-line accessand straight-line access was not formed even in root

formed root canal concentrates stress and has weakcanal preparation by PTN. It was shown that proper

fracture resistance19）. The results showed that end-root canal preparation is possible. Since the PTN X2 fileodontic treated teeth using MI Endo were less suscepti-

has a large taper, flare of the upper one-third of theble to cervical fracture. Previous studies have sug-root canal is formed even in the no straight-line accessgested that the preservation of orifice dentin may begroup（C）when using X2, similar to the straight-line

correlated with long-term maintenance of occlusal func-

access formation, at the apex. It was also shown that14, 15）tion in endodontic treated teeth.

accurate root canal preparation was possible.

Previous research reports have discussed the needIn addition, at 1‒2 mm on the apical side, accurate

for straight-line access in root canal preparation. For

root canal preparation was possible in both the TN andstraight-line access unformed root canals, it is likely to

PTN groups regardless of the presence or absence ofbe difficult to limit the operation of instruments andstraight-line access, and the median root canal value

directly reach a direct view due to the reduction of thewas also the same.

surgical field at the start of root canal preparation.

This result shows that both TN and PTN files have a

Inadequate morphological following ability may result.

6％ taper up to 3 mm on the apical side, TN（apical 0, 1,

The present study analyzed the relationship between

2, 3 mm＝ #26, #32, #38, #44）and PTN（apical 0, 1, 2,

the final canal morphology by Ni-Ti preparation and

3 mm＝ #25, #31, #37, #43）. The fact that the file diam-straight-line access, and accurate root canal preparation

eters were the same may have had an effect. On thewas possible even with MI Endo, which reduces theother hand, the displacement on the inner side of 3, 5

risk of root canal fracture. The 30° curvature of theand 8 mm significantly increased, showing a tendency

curved root canal model used in this study was selectedof straightening on the inner side of the curved root

because it is easy to obtain the effect of reduced curva-

canal in the PTN straight-line access group（D）. Thisture due to straight-line access.

result is considered to be due to the difference in flexi-

Furthermore, the TN used in this study was devel-bility and file taper（4 mm or more from the apex）

oped with the concept of MI Endo, which maintains abetween the PTN file and the TN file. Peters et al.

high occlusal strength of endodontic treated teeth byreported that TN was more fatigue resistant andforming a root canal with minimally invasive shaping.

showed significantly more predictable torque and

The results showed that TN enables proper root canalthreading-in force compared with PTN24）.

preparation without straight-line access. The increase

It was shown that root canal preparation by PTN

in root canal width and the median displacement in TNthat formed the straight-line access, which was consid-

canal preparation were measured. Even when compar-

ered to be essential for maintaining the anatomical root

ing the no straight-line access group（A）and thecanal morphology, straightened the curved root canalstraight-line access group（B）, no significant difference

on the inner side.

was observed between the increase in root canal width

The PTN has two files, X1 and X2, which can obtainand the median displacement using TN. Accurate canal

the final root canal morphology that enables sufficient

preparation was shown to require no straight-line

root canal cleaning and root canal filling, and at theaccess.

same time, the elasticity of the file itself is improved.

On the other hand, a comparison of the PTN with the

The files are easy to use and are expected to reduceno straight-line access group（C）and the straight-line

accidents, and are used clinically. The PTN files areDec, 2021

Canal Transportation by Straight Line Access75

characterized by having a rectangular cross section andalways cutting at only two points in contact with theConclusion

root canal wall in order to reduce the bite into the wall.
Therefore, PTN is the most frequently used file system

Root canal preparation by TN was shown to maintainin Europe and the United States because there are few

the original anatomical root canal morphology without

accidents during root canal preparation and accurate

straight-line access. The minimally invasive shaping

root canal preparation is possible even in curved root

type TN developed based on the concept of MI Endocanals. The TN file system was constructed with a sim-enables accurate root canal preparation and reduces

ilar system based on PTN, and was designed to have a

the amount of root canal displacement after root canal

final root canal morphology that can be filled with root

preparation due to the shape and flexibility of the file.

canals using two files, Small and Prime. However, since
there is no straight-line access, the root canal following

Acknowledgment

ability of the apical curved root canal may be inade-We thank Ashleigh Cooper, PhD, from Edanz（https://jp.edanz.com/ac）for editing a draft of this manuscript. This
study did not involve human participants or animals.

quate due to the restricted instrument operation in thelimited field. The results of this study showed that
there is no need for straight-line access, which has been
considered essential for accurate root canal preparation.

There are no conflicts of interest related to this study.

Insufficient flare preparation in the upper part of theroot canal when straight-line access is not formed suggests the difficulty of root canal cleaning and the effectof insufficient pressurization of root canal obturation.
However, for root canal preparation by minimally invasive shaping type TN, it is recommended to use a dedicated needle with an improved tip for root canal cleaning and pressure root canal obturation or single-pointroot canal obturation with a dedicated gutter core byroot canal obturation. In recent years, a bioceramicsealer has been developed for root canal obturation, andhas been reported to have excellent biological properties such as hard tissue forming ability in addition tobiocompatibility and non-cytotoxicity32）. In particular,
bioceramic sealers are expected to have a higher
tight-sealing chain than other sealers by forming hardtissue at the dentin interface and apical foramen. Currently, the root canal obturation method uses multiple
chemically and physically stable gutta-percha cones totightly seal the root canal by lateral and vertical obturation with heated gutta-percha. The obturation method
has been established and good clinical results have beenreported. However, the single-point root canal filling
method33） using a bioceramic sealer capable of root
canal preparation and hard tissue formation, which
reduces the risk of root canal fracture due to minimallyinvasive cutting type TN, will follow the flow of endodontic treatment in the future. The possibility of shifting to MI Endo was shown.

References
1）Fransson H, Dawson VS, Frisk F, Bjørndal L, EndoReCo
O, Kvist T. Survival of root-filled teeth in the Swedish
adult population. J Endod 2016; 42: 216‒220.
2）Aquilino SA, Caplan DJ. Relationship between crownplacement and the survival of endodontically treatedteeth. J Prosthet Dent 2002; 87: 256‒263.
3）Pratt I, Aminoshariae A, Montagnese TA, Williams KA,
Khalighinejad N, Mickel A. Eight-year retrospective
study of the critical time lapse between root canal completion and crown placement: its influence on the survival of endodontically treated teeth. J Endod 2016; 42:
1598‒1603.
4）Seo DG, Yi YA, Shin SJ, Park JW. Analysis of factors
associated with cracked teeth. J Endod 2012; 38: 288‒
292.
5）Liu R, Kaiwar A, Shemesh H, Wesselink PR, Hou B, Wu
MK. Incidence of apical root cracks and apical dentinal
detachments after canal preparation with hand androtary files at different instrumentation lengths. J
Endod 2013; 39: 129‒132.
6）Fain WD. Tooth fractures and cracks. J Am Dent Assoc
2021; 152: 423‒424.
7）Krishan R, Paqué F, Ossareh A, Kishen A, Dao T, Friedman S. Impacts of conservative endodontic cavity on
root canal instrumentation efficacy and resistance tofracture assessed in incisors, premolars, and molars. J
Endod 2014; 40: 1160‒1166.
8）Moore B, Verdelis K, Kishen A, Dao T, Friedman S.
Impacts of contracted endodontic cavities on instrumentation efficacy and biomechanical responses in max-

76
illary molars. J Endod 2016; 42: 1779‒1783.
9）Goerig AC, Michelich RJ, Schultz HH. Instrumentation ofroot canals in molars using the step-down technique. J
Endod 1982; 8: 550‒554.
10）Swindle RB, Neaverth EJ, Pantera EA Jr, Ringle RD.
Effect of coronal-radicular flaring on apical transportation. J Endod 1991; 17: 147‒149.
11）Torabinejad M. Passive step-back technique. Oral Surg
Oral Med Oral Pathol 1994; 77: 398‒401.
12）Lang H, Korkmaz Y, Schneider K, Raab WH. Impact ofendodontic treatments on the rigidity of the root. J Dent
Res 2006; 85: 364‒368.
13）Tang W, Wu Y, Smales RJ. Identifying and reducing
risks for potential fractures in endodontically treatedteeth. J Endod 2010; 36: 609‒617.
14）Pierrisnard L, Bohino F, Renault P, Barquins M. Corono-radicular reconstruction of pulpless teeth: a mechanical study using finite element analysis. J Prosthet Dent
2002; 88: 442‒448.
15）Clark D, Khademi JA. Case studies in modern molarendodontic access and directed dentin conservation. Dent
Clin North Am 2010; 54: 275‒289.
16）Tang W, Wu Y, Smales RJ. Identifying and reducing
risks for potential fractures in endodontically treatedteeth. J Endod 2010; 36: 609‒617.
17）Reeh ES, Messer HH, Douglas WH. Reduction in tooth
stiffness as a result of endodontic and restorative procedures. J Endod 1989; 15: 512‒516.
18）Pradeep P, Kumar VS, Bantwal SR, Gulati GS. Fracture
strength of endodontically treated premolars: an in-vitro
evaluation. J Int Oral Health 2013; 5: 9‒17.
19）Yuan K, Niu C, Xie Q, Jiang W, Gao L, Huang Z, Ma R.
Comparative evaluation of the impact of minimally invasive preparation vs. conventional straight-line preparation on tooth biomechanics: a finite element analysis.
Eur J Oral Sci 2016; 124: 591‒596.
20）Burllein S, Schafer E. Minimally invasive endodontics.
Quintessence Int 2015; 46: 119‒124.
21）Murdoch-Kinch CA, Mclean ME. Minimally invasive
dentistry. J Am Dent Assoc 2003; 134: 87‒95.
22）Gutamann JL. Minimally invasive dentistry（Endodontics）. J Conserv Dent 2013; 16: 282‒283.
23）Kenneth MH, Stephen C. Cohen’s pathways of the pulp.

ODEP Vol. 1, No. 1
10th ed. St. Louis: Elsevier, 2011; 137‒138.
24）Peters OA, Arias A, Choi A. Mechanical properties of anovel nickel-titanium root canal instrument: Stationary
and dynamic tests. J Endod 2020; 46: 994‒1001.
25）Arias A, Singh R, Peters OA. Torque and force induced
by ProTaper Universal and ProTaper Next during
shaping of large and small root canals in extracted
teeth. J Endod 2014; 40: 973‒976.
26）Nguyen HH, Fong H, Paranjpe A, Flake NM, Jonson JD,
Peters OA. Evaluation of the resistance to cyclic fatigue
among ProTaper Next, ProTaper Universal, and Vortex
Blue rotary instruments. J Endod 2014; 40: 1190‒1193.
27）Shim KS, Oh S, Kum K, Kim YC, Jee KK, Chang SW.
Mechanical and metallurgical properties of various nickel-titanium rotary instruments. Biomed Res Int 2017;2017: 4528601 doi: 10.1155/2017/4528601
28）Chen G, Fan W, Mishra S, El-Atem A, Schuetz MA, Xiao
Y. Tooth fracture risk analysis based on a new finite
element dental structure models using micro-CT data.
Comput Biol Med 2012; 42: 957‒963.
29）Al-Omiri MK, Rayyan MR, Abu-Hammad O. Stress analysis of endodontically treated teeth restored with
post-retained crowns: a finite element analysis study. J
Am Dent Assoc 2011; 142: 289‒300.
30）Belli S, Eraslan O, Eraslan O, Eskitascioglu G. Effect of
restoration technique on stress distribution in roots
with flared canals: an FEA study. J Adhes Dent 2014;
16: 185‒191.
31）Sathorn C, Palamara JE, Palamara D, Messer HH. Effectof root canal size and external root surface morphology
on fracture susceptibility and pattern: a finite element
analysis. J Endod 2005; 31: 288‒292.
32）Sanz JL, Rodríguez-Lozano FJ, Llena C, Sauro S, Forner
L. Bioactivity of bioceramic materials used in the dentin-pulp complex therapy: A systematic review. Materials 2019; 12: 1015. doi: 10.3390/ma12071015
33）Yu YH, Kushnir L, Kohli M, Karabucak B. Comparing
the incidence of postoperative pain after root canal filling with warm vertical obturation with resin-based
sealer and sealer-based obturation with calcium silicate-based sealer: a prospective clinical trial. Clin Oral
Investig 2021; 25: 5033‒5042.

Dec, 2021

77

Original Article

ODEP 1（1）：77～87，2021

Study on Evaluation of Cavity Preparation Using
Non-contact High-speed 3-D Shape Measuring Device
Saeko OKUMURA, Hiroaki TANIMOTO, Kenzo YASUO, Naohiro IWATA,
Kazushi YOSHIKAWA and Kazuyo YAMAMOTO
Department of Operative Dentistry, Osaka Dental University

Abstract
Introduction: In dental education, it has long been essential to master cavity preparation techniques bytraining with models, both during basic and clinical training before college graduation as well as during clinical practice as dentists. However, the instruction given is subjective depending on the instructor, with variations in instruction contents and evaluations, and it has long been pointed that quantification is difficult.
Effective and efficient education and instruction as well as reproducible and objective evaluations are essential. Therefore, this study examined the objective evaluation and grading of Class Ⅱ metal inlay cavity by acomputer system using a non-contact high-speed 3-D shape measuring device.
Subjects and methods: Ten dental residents at the Department of Operative Dentistry, Osaka Dental University Hospital were recruited as subjects. A jaw model containing an artificial tooth for training was used
to conduct cavity preparation for a mesial Class Ⅱ metal inlay on the mandibular right first molar tooth
within 30 minutes. After preparation, an evaluation was conducted by this system and by nine instructors at
the Department of Operative Dentistry with 7 to 28 years of clinical experience. A free description column
for the evaluation and instruction items was provided as part of the evaluation by instructors. Feedback wasgiven based on the results, and a total of five sessions of cavity preparation were conducted at two-monthintervals. Pearson’s correlation coefficient was used in the correlation analysis of the evaluation results.
Results: Reanalysis was conducted by excluding the seven points which had the largest deductions in theevaluation using the measuring device. As a consequence, a significant positive correlation with the numberof preparations was observed. There was also a significant positive correlation with the evaluation by
instructors. A comparison of the evaluations by the measuring device and by the instructors showed a significant positive correlation.
Conclusion: The measuring device can consistently and objectively evaluate Class Ⅱ metal inlay cavity
preparation equivalent to evaluations by multiple instructors with long clinical experience. It was also shownthat a curriculum using evaluations by the measuring device and instructors effectively improved the skillsof the dental residents. Incorporating evaluation by the measuring device may be an effective method for
reducing the burden on instructors and length of the instruction period.
Key words: objective evaluation, cavity preparation, dental education

Corresponding author: Saeko OKUMURA, Department of Operative Dentistry, Osaka Dental University, 1‒5‒17, Otemae,
Chuo-ku, Osaka 540‒0008, Japan
TEL: ＋81‒6‒6910‒1087, FAX: ＋81‒6‒6910‒1052, E-mail: okumura-s@cc.osaka-dent.ac.jp
Received for Publication: September 10, 2021/Accepted for Publication: October 15, 2021
DOI: 10.11471/odep.2021‒011

ODEP Vol. 1, No. 1

78

tistry, Osaka Dental University Hospital, which is acontrolled postgraduate education institution.

Introduction

2 ．Methods
In the field of dental education, it has long been

A jaw model（D16FE-500H（GSF）-MF, Nissin Dental

essential for students to master tooth-cutting tech-

Products Inc, Kyoto）, in which an artificial tooth forniques. In particular, the technique used to prepare atraining（A5SA-500#46, Nissin Dental Products Inc.）

cavity for metal inlay, which is an indirect repair

with special processing on melamine was inserted as

method, greatly affects the prognosis after treatment.

the mandibular right first molar tooth, was attached toConventionally, students learn the basic skills of metal

the simulator（DR-H Ni: MO, Nissin Dental Products

inlay cavity preparation by repeatedly training using

Inc.）（Figs. 1 and 2）. The task was to form a mesial

mannequins and jaw models during undergraduate

Class Ⅱ metal inlay cavity（within 30 minutes）. A

basic and clinical training before college graduation as

period of two minutes to visually observe the example

well as during clinical practice as residents. At present,

tooth（Fig. 3）was given, and the subjects were notthere are no other methods of evaluation or instruction;

allowed to check the example during cavity prepara-

both of these approaches are subjective, depending ontion. As cutting tools, diamond burs（#401, #301, #F215,

the instructor or medical advisor. An instructor with

#201, SHOFU Inc, Kyoto）, and steel burs（UA, No. 701,

experience and skills who has mastered cavity prepara-

No. 702, No. 703 Steel, Beldenta Supply Inc, Switzer-

tion techniques is thus required for evaluation andland）were used（Fig. 4）. The formed cavity was mea-

instruction. However, students often become confused

sured and evaluated with the 3-D shape measuring

due to differences in the instructions given by eachdevice（hereafter,“evaluation by measuring device”）,

instructor, as well as variations in contents and evalua-

and was evaluated by nine instructors belonging to thetions. In addition, providing instruction in the clinical

Department of Operative Dentistry with 7 to 28 years

field requires much time and effort, sometimes result-of clinical experience（hereafter,“evaluation by instruc-

ing in failure to provide sufficient instruction1）.

tors”）. A column for free description on evaluation andCavity preparation techniques are often evaluatedinstruction items was provided as part of the evaluation

based on subjective observation of outline and depth of

by instructors（Table 1）. Feedback was given to eachthe cavity by the instructor. Even the same instructor

student based on the results after evaluation. Cavity

may sometimes give different evaluation results2‒5）since

preparation was conducted at two-month intervals in

6）quantification is difficult , and the lack of objectivity inthe curriculum, with a total of five cavity preparation

evaluation methods has been pointed out in the past6,7）.

sessions, followed by measurement, evaluation, grading

Some have reported that effective and efficient educa-

and feedback. Informed consent was orally given totion and instruction as well as reproducible evaluations

participants in the research and the freedom to refuse

2,3,8‒10）. In addition, with the use of 3-D shapeparticipation was guaranteed. Pearson’s correlation

measuring devices, it is now possible to measure thecoefficient was used in the correlation analysis between

are essential

1,11,12）

data on cavity preparation in a short period.

This study examined the consistency of objective

the evaluation by measuring device and the evaluation
by instructors.

evaluations by instructors by using a computer system

3 ．Grading evaluation methods

featuring non-contact high-speed 3-D shape measure-1 ）Cavity preparation evaluation system and mea-

ment to perform objective evaluations and grading. The
usefulness and practicality of a curriculum using thissystem was also examined.

surement principle
The system used in this study consisted of a 3-D
shape measuring device（J. Morita Mfg. Corp, Kyoto;
hereafter,“SURFLACER”）（Figs. 5 and 6）, a personal

Subjects and Methods

computer（Latitude E5430, DELL, Tokyo）
equipped withthe measuring device control software and cavity

1 ．Subjects

preparation skill evaluation software, and a printer（LP-The subjects were ten dental residents undergoing

S520, EPSON, Tokyo）1）. The system measured theclinical training at the Department of Operative Den-formed cavity using SURFLACER. It superimposed theDec, 2021

Fig. 1

The Device can Objectively Evaluate Metal Inlay Cavities and are Consistent

Artificial tooth for training

Fig. 2

Simulator, jaw model

Table 1

Fig. 3

79

Example tooth

Instructor evaluation items and allocation of points

Cavity outline

Allocationof points

1．Size（width）of the cavity outline□ Too large（too wide）□ Rather large（wide）□ Appropriate□ Rather small（narrow）□ Too small（too narrow）
Fig. 4

Burs that were used

10
20
35
25
15

2．Is the cusp ridge retained?

executed evaluation and grading on the computer using□ Retained□ Somewhat retained□ Difficult to say□ Retention rather failed□ Not retained

the different date.

3．Does the cavity outline include the pit and fissure?

example tooth cavity shape data that was registered inadvance and the shape data for the formed cavity, andThe principle of measurement was as follows. The
laser was irradiated on the formed tooth attached tothe special jig and the slit light of the laser wasreceived by two CMOS cameras. The 3-D coordinate
values on the specimen were obtained by triangulation
measurement using the slit-ray projection method.
Computer calculations were made based on these 3-D
data to convert them into an image1,13）. The basic triangulation measurement principle using the slit-ray projection method is shown in Fig. 7. The optical axis of
the laser projection part was positioned at angle θ tothe optical axis of the CMOS camera detection part.
Since image formation point P’ on the horizontal imageof the CMOS camera changes when the position of theintersection point P moves from camera center O, it ispossible to calculate height Z（height of actual object）
as displacement Z from the camera center by reading
the position of image point P’.

□ Included□ Somewhat included□ Difficult to say□ Rather not included□ Not included
Cavity depth

30
25
18
12
5

30
25
18
12
5
Allocationof points

4．Cavity bottom dept
□ Too shallow□ Somewhat shallow□ Appropriate□ Somewhat deep
□ Too deepFree description column

35
40
95
40
20

ODEP Vol. 1, No. 1

80aRelative deduction

Relative deduction
Deduction ratio

0

3

1

Absolute deduction
Deduction value

Fig. 8Fig. 5

SURFLACER

10

10

0

0.5

1

Deduction value

10

Evaluation map for outside and inside the cavity
a. Outside the cavity b. Inside the cavity

Cavity depth evaluation map

Insufficient cutting

Attachment part of SURFLACER
for artificial tooth for training

Deduction ratio
Absolute deduction

Table 2

Fig. 6b

Excessive cutting

Shallowness（mm）

Deduction
ratio

Depth
（mm）

Deduction
ratio

0.00‒0.10
0.10‒0.20
0.20‒0.30
0.30‒0.40
0.40‒0.50
0.50‒

0.00
3.00
1.00
1.00
1.00
0.00

0.00‒0.10
0.10‒0.20
0.20‒0.30
0.30‒0.40
0.40‒0.50
0.50‒

0.00
0.25
3.00
1.00
1.00
0.00

2 ）Setting the criteria for evaluation with the measuring device
A cavity that satisfied the requirements for a metal
inlay cavity14,15） was formed as the example tooth. It is

Projection elements
Laser beam（Slit-ray source）
Detection elements
CMOS camera

Polygon mirror

O’P’

Detection elements
CMOS camera
O’Fig. 7in advance and set up the evaluation map based on theincorporated data in order to execute evaluation by themeasuring device. The evaluation map is divided into
the outline and depth of cavity. The cavity outline was
set up by dividing it into maps to evaluate and grade
the outer or inner side of the margin of the example

L2

tooth. The area outside the margin was considered as
Displacement Z

Measurement range（Height : Z）

L1

Point laser

P’

necessary to incorporate the data on the example tooth

P
θ θ

O
Center of camera

Slit‒ray projection method measurement principles

the excessive cutting area, and the area inside the margin as the insufficient cutting area, and the ratio of
deduction and locations that could not be cut or that
could not be left uncut were set up（Fig. 8, Table 2）.
Regarding cavity depth, dental pulp exposure is a contraindication in clinical practice. Excessive cutting wastherefore set with a larger deduction ratio than insufficient cutting.

Dec, 2021

The Device can Objectively Evaluate Metal Inlay Cavities and are Consistent
Table 381

Score calculation formula for SURFLACER

1/2
Cavity outline evaluation score O＝（OE×OI）

Evaluation score for excessive cutting OE＝100－excessive cutting deduction value

Evaluation score for insufficient cutting OI＝100－insufficient cutting deduction value
1/2
Cavity depth evaluation score D＝（DE×DI）

Evaluation score for excessive cutting DE＝100－excessive cutting deduction value
Evaluation score for insufficient cutting DI＝100－insufficient cutting deduction value
1/2
Total evaluation score＝（O×D）

Score calculation formula for instructor evaluation

based on the results of a preliminary experiment. The

Cavity outline evaluation score＝（evaluation item 1＋2＋3）
Cavity depth evaluation score＝evaluation item 4
1/2
Total evaluation score＝（outline score×depth score）

highest and lowest scores by the nine instructors wereTable 4also excluded and the average of the other instructors
was used as the score.
5 ）Feedback method
Feedback was given to the dental residents using the3 ）Setting the evaluation criteria for the evaluation
by instructors

results of evaluation by measuring device（Figs. 9 and
10）and the results of evaluation by instructors. The

The evaluation by instructors was made using theparts in the evaluation by measuring device that aresame example tooth used for the evaluation by measur-

marked in red indicate excessive cutting, and those ining device. The teeth formed by dental residents were

blue indicate insufficient cutting. The results allow visu-

assigned codes A through J so that the individuals

alization into fine parts. The average score of the nine

could not be identified.

instructors was indicated for the evaluation by instruc-4 ）Score calculation in evaluation by measuring
device and evaluation by instructors

tors, and the comments in the free description column
were presented along with the score.

Table 3 shows the score calculation formula for SURFLACER in the evaluation by measuring device. In

Results

addition, points for the outline and points for the depthof cavity were both calculated in the deduction method.

The total score results of the evaluation by measur-

Tables 1 and 4 show the evaluation items and allocationing device and the evaluation by instructors are shownof points in the evaluation by instructors, and the scorein Table 5.

calculation formula. The average score resulting from

1 ．Objective evaluation by measuring device

five measurements and gradings on the example toothThe results of correlation analysis showed no signifi-

after setting the criteria for the evaluation by measur-cant difference between the number of preparations

ing device was 95 points. Based on this, the point alloca-

and evaluation by measuring device（Fig. 11）（NS）. The

tion was set so that the highest score would be 95

average score was 60 points and the highest score 85

points for the third item in cavity outline and the firstpoints. In the results, the largest deduction（ －98

item in cavity depth. In accordance with clinical prac-

points）occurred at seven points（A-3, C-2, -3 and -5, F-3,

tice in a similar fashion to the setting for the evaluation

G-3 and -4）
due to the locations at which cutting or leav-by measuring device, both cavity outline and cavity

ing uncut was a contraindication, resulting in especiallydepth were set so that a lower score was given when

low scores（Table 5）. A breakdown of the scores at

cutting was excessive. Settings were also made to mini-these seven points is shown in Table 6. Of the sevenmize the variation in scores among the instructors

points, excessive cutting in cavity depth was consideredODEP Vol. 1, No. 1

82

Prepared Cavity Evaluation Result

Prepared Cavity Evaluation Result
Sample1
Tooth ID: #46
Cavity ID:

Tooth name:
Cavity name : ClassⅡ

Sample2
Tooth ID: #46
Cavity ID:

Tooth name:
Cavity name : ClassⅡ

Cavity evaluation

Vertical section

Cavity evaluation

Vertical section

Horizontal section

Master Cavity

Horizontal section

Master Cavity

Student

Student

MasterMasterExcessive
cutting

Excessive insuﬃcientcutting
cutting

Insuﬃcientcutting

Total score 70
Outline form score 72
Demerit score for excessive cutting 29
Demerit score for insuﬃcient cutting 27

Fig. 9Depth score 69
Demerit score for excessive cutting 27
Demerit score for insuﬃcient cutting 35

Measuring device evaluation results,
Example（1）Correlation analysis showed a significant positive cor-

80
Score

Measuring device evaluation results,
Example（2）2 ．Objective evaluation by instructors

90

relation between the number of preparations and evalu-

70

ation by instructors（Fig. 13）（r＝0.30, p＜0.05）
. The

60

evaluation by instructors tended to be higher in both

50

average score and highest score.

40

3 ．Comparison between evaluation by measuring

30

Fig. 11

Fig. 10

Depth score 72
Demerit score for excessive cutting 11
Demerit score for insuﬃcient cutting 41（r＝0.32, p＜0.05）.

100

20

Total score 76
Outline form score 80
Demerit score for excessive cutting 30
Demerit score for insuﬃcient cutting 9

1

2

3

4

NS
5 Session

Measuring device evaluation score（by
preparation session）

device and evaluation by instructors
A scatter diagram of the evaluation by measuring
device and the evaluation by instructors is shown in
Fig. 14. Correlation analysis showed a significant positive correlation between the two evaluations（r＝0.41,
p＜0.01）.

a contraindication at six points and excessive cutting incavity outline was a contraindication at one point. Cor-

Discussion

relation analysis was therefore conducted by excluding
the above seven points, and the result showed a signifi-

For improving clinical dental skills, it is necessary tocant positive correlation between the number of prepa-

master the procedures in addition to learning therations and evaluation by measuring device（Fig. 12）

knowledge in order to conduct treatments. In dental

Dec, 2021
Table 5

The Device can Objectively Evaluate Metal Inlay Cavities and are Consistent
Measuring device/instructor evaluation

100

score results

90
80

1

2

3

4

5

A

DS
IS

52
57

66
70

30
42

70
79

70
69

B

DS
IS

75
74

57
56

68
75

69
75

59
64

C

DS
IS

60
49

25
67

28
84

70
92

34
50

D

DS
IS

36
46

64
54

65
55

60
56

59
82

E

DS
IS

63
76

66
78

55
60

71
92

60
74

F

DS
IS

71
81

53
69

35
58

70
88

85
93

100

G

DS
IS

56
58

59
85

27
67

29
92

70
89

80

H

DS
IS

58
46

66
88

46
55

76
86

63
59

I

DS
IS

73
71

66
70

72
70

67
56

67
61

J

DS
IS

58
61

73
62

82
92

58
73

78
85

（Session）

Table 6

A‒3
C‒2
C‒3
C‒5
F‒3
G‒3
G‒4

Score breakdown

T

O

OE

OI

D

DE

DI

30
25
28
34
35
27
29

65
62
13
83
87
76
78

91
45
2
75
98
91
90

46
85
81
91
78
62
69

14
10
60
14
14
9
11

2
2
37
2
2
2
2

100
47
98
100
100
45
61

T：Total evaluation score, O：Cavity outline evaluation
score, OE：Evaluation score for excessive cutting, OI：
Evaluation score for insufficient cutting, D：Cavity depth
evaluation score, DE：Evaluation score for excessive cutting, DI：Evaluation score for insufficient cutting

Score

70
60
50
40

r＝0.32 p＜0.05

30
20
Fig. 12

1

2

3

4

5 Session

Measuring device evaluation score（by
preparation session, with deletion ofseven points）

Score

90
70

r＝0.30 p＜0.05

60
50
40
30
20

Fig. 13

1

2

3

4

5 Session

Instructor evaluation score（by preparation session）

100

Measuring device evaluation score

DS：Measuring device evaluation score, IS：Instructor
evaluation score

83

90
80
70
60
r＝0.41 p＜0.01

50
40
30
20
20

Fig. 1430

40 50 60 70 80 90
Instructor evaluation score

100

Correlation diagram on measuring
device evaluation score and instructor evaluation scorecare, precise work must be conducted inside the con-within the oral cavity of the patient, it is important tofined space of the oral cavity. To acquire the skills to

repeatedly perform skill training in basic education andconduct diagnosis and treatment safely and precisely

training. It is therefore necessary to evaluate skills

ODEP Vol. 1, No. 1

84
Cavity width
CameraLaser slit

Cavity evaluation

Vertical section

Horizontal section

Master Cavity

CameraMeasurement limit
depth

Parallaxangle
60
degrees

ApproxFig. 15

Invisible area
Student

Table 7

MasterMeasurable depth to cavity width

Cavity width（mm）
0.5
Measurable depth（mm） 0.45

1.0
0.90

1.5
1.35

2.0
1.80

2.5
2.25

Excessive
cutting

Insuﬃcientcutting

Measuring device evaluation score
Total score 28
Outline form score 13
Demerit score for excessive cutting 98
Demerit score for insuﬃcient cutting 19

Depth score 60
Demerit score for excessive cutting 63
Demerit score for insuﬃcient cutting 2

Instructor evaluation score
Total score 84
Outline form score 74
Depth score 95

Fig. 17

Results of evaluation by measuring device
and instructors（C‒3）

having sufficient practical experience as it is becoming
increasingly difficult to gain sufficient clinical skills at
outpatient hospitals. Therefore, this study comparedobjective evaluation by measuring device and objective

Example of invisiblearea display○：Invisible area

Fig. 16

evaluation by multiple instructors in cavity preparation
for a Class Ⅱ metal inlay, and examined the usefulness
and practicality of a skill education curriculum that
adopted this system.

objectively, not based on the subjectivity of the instruc16‒23）

This study recruited dental residents who were

. Unlike

undergoing clinical practice at the Department of Oper-knowledge that can be learned in the classroom, practi-

ative Dentistry, Osaka Dental University Hospital. The

cal experience is essential for mastering the proce-skills of a dental resident improve by both experience

dures. This is one of the reasons why clinical training isin diagnosis and treatment and self-improvement using

included as a requirement in curriculums for dental

jaw models on which they work during spare time fromschool students. Universities that teach dentistry thus

diagnosis and treatment. Accordingly, medical advisors

have university hospitals where students can receive

assign patients to suit the skills of the dental resident.

clinical training. However, the number of patients who

The study was therefore conducted within the extent

participate in clinical training has declined dramaticallythat would not interfere with their diagnosis and treat-

in recent years due to changes in patients’ attitudes

ment at two-month intervals. No significant difference

toward medicine, etc., and many dentistry students facein correlation was observed in total scores of evaluation

the risk of graduating and becoming dentists without

by measuring device. It is considered that the maxi-tor, and various attempts have been made

Dec, 2021

The Device can Objectively Evaluate Metal Inlay Cavities and are Consistent85

mum deduction at seven points（－98 points）affected

done.”Instructors tended to give a high evaluation as

the correlation results greatly（Table 5）. The cut onlong as the tooth was formed with a good balance, evenlocations uncut considered as contraindication werewhen parts that were subject to deduction had been

specified in the settings for the measuring device. As acut. While dental residents may feel that evaluations

result, large deductions occurred. As a significant posi-done by measuring device under certain conditions aretive correlation was observed by excluding these seven

severe, it is considered better to evaluate under certain

points, it was found that skills were improved by con-conditions when comparing changes over time. Since

ducting evaluations by measuring device and giving

the evaluations by measuring device and by instructors

feedback over time. It was also found that skills were

showed a significant positive correlation, it was consid-

improved by conducting evaluations by instructors andered that the measuring device can perform evaluations

giving feedback over time, as a significant difference inthat are similar to those by instructors.

correlation coefficient was observed in the evaluations

It is possible to apply SURFLACER, which was usedby instructors. Based on these findings, this curriculum,in this study, in basic training and clinical training

which incorporates evaluation by measuring device andbefore graduation in addition to the training of dental

evaluation by instructors, is expected to be effective forresidents. By incorporating phantom training using thisimproving the skills for cavity preparation.

system, it is expected that students can experience

Evaluation of cavity preparation with measuring

more complete training as the limited skill training

devices has been used to ensure fairness for students

period can be utilized more effectively and the instruc-and make feedback more objective2,7,24‒28）. Meanwhile,

tor can spend more time in instructing individual stu-

evaluations by instructors have also been examined,

dents. Furthermore, SURFLACER can be operated byincluding whether importance should be placed on thethe students or dental residents themselves, and theycavity outline or depth, and what ratio of the two would

can receive instant self-feedback29）. It is also considered

be best1）. Variations in evaluations among instructorsthat the system will help students study better on theirwere also observed in some of the formed teeth in thisown, as it can be used to visualize even the points ofstudy. On the other hand, evaluation by measuring

the formed tooth that are difficult to see with the eye,device is possible under certain conditions, as a detailed

and make it easier to understand visually with coloring.

evaluation map can be set up and differences that can-Since reports have been published on using the system

not be judged by the human eye can be evaluated.

for training on abutment tooth preparation, caries

However, there are areas that cannot be seen by theremoval and pulp chamber opening other than cavity

measuring device（Fig. 15）, and the measurable depth

preparation2,29,30）, it is considered that SURFLACER isvaries depending on the cavity width（Table 7）. If there

an effective new method of skill training. Because theare invisible areas within the cavity, it is impossible toevaluation map on the example tooth can be freely set,evaluate the cavity depth in such areas, even thoughanother advantage of this system is that the severity of

the cavity outline can be evaluated. In such cases, theevaluation can be specified to suit the skill level of thecavity depth is evaluated based on the points in prox-

student. It can be set to an easy level for basic training

imity to the invisible area that can be measured. Thatof students who have little experience in cavity prepa-

is, evaluation by an instructor is necessary for 3-D

ration, and to a harder level as they gain experience

shape measurement using laser light, as the occurrence

from hospital training on the way to becoming dental

of invisible areas is unavoidable. The invisible areas canresidents. It may also be used by dentists as a self-im-

be recognized by experienced people, as they appear asprovement tool for ongoing training.

white areas in the results（Fig. 16）.
At C-3, which had the largest deduction in the evalu-

Conclusion

ation by measuring device, the evaluation by instructors varied greatly with a tendency for high evalua-

1．The non-contact high-speed 3-D shape measuring

tions（Fig. 17）. Comments in the free description column

device can objectively evaluate Class Ⅱ metal inlay

in the instructor evaluation included“it was well-cavities, and can execute evaluations that are consistent

formed,”“slightly too large on the lingual side, but well-with evaluations by multiple instructors with rich clini-ODEP Vol. 1, No. 1

86

cal experience.

Japanese Society of the General Dentistry 2019; 11: 58‒

2．A curriculum that includes evaluation by measuring device and evaluation by instructors is effective for
improving the skills of dental residents.
3．The incorporation of evaluation by measuring
device may reduce the burden on instructors and the
instruction period.
Acknowledgement
We thank the members of the Department of Operative
Dentistry, Osaka Dental University, for their advice andhelp.
This study was supported in part by Osaka Dental University Research Funds（21-02）.
The authors declare no conflicts of interest associatedwith this manuscript.

References
1）Hirata K, Nakashima M, Imura S, Yamamoto K, Sekine
I, Moriwaki Y, Yoshida S. Application of noncontact
high speed 3-D shape measuring system to restorative
dentistry Part 2 Development of cavity preparationassessment system. Jpn J Conserv Dent 1998; 41: 320‒
336.（in Japanese）
2）Yamaguchi T, Hosoya N, Kurachi Y, Yoshida T, Morito
A, Koda S, Yamamoto T, Momoi Y. Objective evaluationof access cavity preparation. Jpn J Conserv Dent 2012;55: 278‒284.（in Japanese）
3）Yasunaga T, Kato S, Takaoka T, Nishi M, Miura M,
Ebisu S. Uniformization of subjective evaluation on class
Ⅰ inlay cavity. DSTS 1996; 91‒98.（in Japanese）
4）Kimoto S, Matsuzawa M, Kubota M, Komatsu T, Yamaguchi M. Evaluation by objective structured clinicalexamination（OSCE）during pre-clinical practice forpediatric dentistry: A study of evaluation results by
instructors and students. Bull Kanagawa Dent Col 2007;
42: 32‒41.（in Japanese）
5）Itoh N, Osawa G, Ogawa T, Kawamura H, Nakaya H.
Evaluation of scaling rootplaning in pre-clinical training
program by objective structured clinical examination―
Results of 6 years―. J Jpn Soc Periodontol 2010; 52:429‒433.（in Japanese）
6）Oka A, Murayama R, Takano R, Furuichi T, Sai K,
Takeuchi Y, Seki K, Furuchi M, Kamimoto A, Masutani
S. A study of cavity preparation training using digital
data obtained by optical impression device. Journal of65.
（in Japanese）

7）Torii M, Yasunaga T, Urano M, Takeshige F, Tsuchitani
Y. Numerical evaluation of prepared cavity and its
application to simulation training. The Japanese Journalof Ergonomics 1995; 31: 101‒104.
（in Japanese）
8）Taguchi Y, Takizawa T, Oka Y, Fujii N. A study of clinical training and evaluation for the preparation of afull-veneer crown. Ann Jpn Prosthodont Soc 2012; 4:
434‒443.
（in Japanese）
9）Quinn F, Keogh P, McDonald A, Hussey D. A pilot study
comparing the effectiveness of conventional training
and visual reality simulation in the skills acquisition ofjunior dental school. Eur J Dent Educ 2003; 7: 13‒19.
10）Quinn F, Keogh P, McDonald A, Hussey D. A study comparing effectiveness of conventional training and virtual
reality simulation in the skills acquisition of junior dental school. Eur J Dent Educ 2003; 7: 164‒169.
11）Hirata K, Nakashima M, Imura S, Yamamoto K, Sekine
I, Moriwaki Y, Yoshida S. Application of non-contact
high speed 3-D shape measuring system to restorative
dentistry Part 1 Accuracy in measuring width anddepth of standard cavity. Jpn J Conserv Dent 1997; 40:287‒293.
（in Japanese）
12）Hirata K, Nakashima M, Imura S, Yamamoto K, Sekine
I, Moriwaki Y, Yoshida S. Application of non-contact
high speed 3-D shape measuring system to restorative
dentistry Part 3 Assessment of prepared cavity bypoints. Jpn J Conserv Dent 1998; 41: 337‒342.（in Japanese）
13）Hirata K. Application of laser measuring system to dental education. JJSLSM 2003; 24: 168.
（in Japanese）
14）Tagami J, Nara Y, Yamamoto K, Saito T. Operative Dentistry 21. 5th ed. Nagasue Shoten: Kyoto; 2017. 112‒118.
（in Japanese）
15）Kitamura C, Morotomi T. Required conditions for cavity
preparation. Senda A, Miyazaki M, Hayashi M, Mukai Y,
Saito T. Operative Dentistry. 7th ed. Ishiyaku Publishers, Inc.: Tokyo; 2019. 114‒122.
（in Japanese）
16）Kimura K. Studies on the development of the systemusing a digital camera for the evaluation of cavity
preparations. J Hiroshima Univ Dent Soc 2004; 36: 24‒
36.
（in Japanese）
17）Ibaraki Y, Haraguchi K, Tsukagoshi S, Odachi T, Nagai
Y, Saito T, Kawakami T, Kobayashi F, Matsuda K.
Rationalization and improving efficiency of dental clinical education using simulation systems. Jpn J Conserv
Dent 2002; 45: 52‒61.
（in Japanese）
18）Kawamoto T. Application of a computer to dental education―Application to the lecture―. JJDEA 1995; 11: 97‒
100.
（in Japanese）
19）Kawamoto M, Kitano T, Kawamoto T, Onchi Y, Kohmi

Dec, 2021

The Device can Objectively Evaluate Metal Inlay Cavities and are Consistent87

T, Inoue M, Narikawa K, Fujii B. Application of 3D

H, Maki K, Inoue M. Method of evaluation of preclinicalcomputer-graphics to dental education Part 1 Develop-

tooth modeling practice in pediatric dentistry and auto-

ment of the trial teaching aids for bevel preparation ofmatic evaluation of 3D shape. Jpn J Ped Dent 2015; 53:

metal inlay cavity. JJDEA 1995; 11: 101‒107.
（in Japanese）
20）Gonda T, Maeda Y, Nokubi T. Multimedia support system for clinical prosthodontic education. The Journal of
Osaka University Dental Society 1997; 42: 148‒153.
（inJapanese）
21）Kurachi Y. Objective evaluation of cavity preparation―Class Ⅱ cavity―. TUDJ 2002; 28: 95‒107.
（in Japanese）
22）Johnson L, Thomas G, Dow S, Stanford C. An initial evaluation of the Iowa Dental Surgical Simulator. J Dent
Educ 2000; 64: 847‒853.
23）Kota K, Ayukawa Y, Iwaku M. An introduction of computer to dental simulation system Part 1 Evaluation
methods for cutting pressure and prepared cavities. Jpn
J Conserv Dent 1994; 37: 152. P‒62.（in Japanese）
24）Yasukawa Y. The effectiveness of cavity preparation
training using a virtual reality simulation system withor without feedback. J Stomatol Soc Jpn 2009; 76: 73‒
80.
（in Japanese）
25）Yamada S, Ono Y, Sugiyama T, Watari M, Asari J,
Mayahara M, Yanagisawa W, Futaki K, Sato M, Suzuki

471‒476.
（in Japanese）

26）Wierinck E, Puttemans V, Swinnen S, Van Steenberghe
D. Effect of augmented visual feedback from a virtual
reality simulation system on manual dexterity training.
Eur J Dent Educ 2005; 9: 10‒16.
27）Welk A, Maggio MP, Simon JF, Scarbecz M, Harrison
JA, Wicks RA, Gilpatrick RO. Computer-assisted learning and simulation lab with 40 DentSim units. Int J
Comput Dent 2008; 11: 17‒40.
28）Abe T, Okuyama Y, Kasahara S, Kimura K. Education
effect on tooth preparation of visual feedback using
computer graphics. Ann Jpn Prosthodont Soc 2009; 1:
123‒129.
（in Japanese）
29）Huiru Z, Shufeng J, Jianping S, Yanmei D. A cavity
preparation evaluation system in the skill assessment ofdental students. J Dent Educ 2016; 80: 930‒937.
30）Wakamatsu N, Okano T, Kondo T, Hasegawa S, Iinuma
M. Objective evaluation of abutment tooth preparation
for preformed metal crowns for primary molars in Japanese preclinical training. Pediatr Dent J 2020; 30: 92‒
98.

ODEP Vol. 1, No. 1

88

Original Article

ODEP 1（1）：88～96，2021

Effects of Universal Adhesives on Dentin Adhesive Performanceand Acid-base Resistance
Ryuta ANDO1, Takaaki SATO1, Naoko MATSUI1,
Masaomi IKEDA3, Noriko HIRAISHI1, Tomohiro TAKAGAKI1,2,
Toru NIKAIDO1,2, Junji TAGAMI1 and Yasushi SHIMADA1
1

Department of Cariology and Operative Dentistry, Division of Oral Health Sciences, Medical and Dental Sciences Track,
Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University（TMDU）
2

Department of Operative Dentistry, Division of Oral Functional Science and Rehabilitation,
Asahi University, School of Dentistry3

Clinical Oral Science, Department of Oral Health Care Sciences, Tokyo Medical and Dental University（TMDU）

Abstract
Purpose: This study elucidated the dentin-bonding performance of three kinds of universal adhesive using
the bond strength test and morphological observation under a scanning electron microscope（SEM）.
Methods: OptiBond Universal（OPB）, Prime&Bond Universal（PBU）, and Clearfil Universal Bond Quick ER（UBQ）were used. For the microtensile bond strength（μTBS）test, human dentin was ground with #600grit silicon carbide（SiC）paper, then an adhesive and built resin composite were applied. After water storagefor 24 h, bonded teeth were sectioned into beam-shaped specimens. Specimens were divided into three subgroups and exposed to 0, 5,000, or 10,000 thermal cycles. After the μTBS test, statistical differences were
examined by multiple comparisons using t-tests with Bonferroni correction（α＝0.05）. For the acid-baseresistant zone（ABRZ）observation, human molars were sectioned into dentin disks. Dentin surfaces wereground with #600-grit SiC paper. Following the application of the adhesive, a flowable resin composite waslight-cured to make a dentin disk sandwich. After water storage, the acid and base challenges were conducted using a buffered demineralizing solution（pH＝4.5）and 5％ NaOCl（pH＝10.5‒12.9）. Then, the surface
was covered by adhesive resin Super-Bond C&B. The specimens were sectioned to approximately 2 mm
thickness, and polished. The polished surfaces were etched with an argon-ion beam, and gold-sputter coatedfor the morphological analysis of adhesive interface under SEM.
Results: The results of μTBS tests depended on the thermal cycle challenges. After 10,000 cycles, theμTBS of all adhesives were significantly lower than those of the 0 cycle group. After 5,000 cycles, μTBS
results showed different trends among the three groups. ABRZ observation showed that all groups formed
ABRZ. A funnel-shaped erosion was clearly observed in OPB and PBU, but not in UBQ. On the other hand,funnel-shaped erosion was observed in UBQ, but not clearly.
Conclusion: The bond strength test results and the interfacial morphological structure after the acid-basechallenge depended on material components such as hydrophilic amide monomer, HEMA and NaF. The
results of this study suggest that the bonding durability between universal adhesives and dentin depends onmaterial components.
Key words: universal adhesives, dentin bond strength, acid-base resistance zone
Corresponding author: Takaaki SATO, Department of Cariology and Operative Dentistry, Division of Oral Health Sciences,
Medical and Dental Sciences Track, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University（TMDU）, 5‒45, Yushima 1-chome, Bunkyo-ku, Tokyo 113‒8549, Japan
TEL: ＋81‒3‒5803‒5483, FAX: ＋81‒3‒5803‒0195, E-mail: t.sato.ope@tmd.ac.jp
Received for Publication: September 15, 2021/Accepted for Publication: October 25, 2021
DOI: 10.11471/odep.2021‒012

Dec, 2021

Effects of Universal Adhesives on Dentin Adhesive and Acid-base Resistance89

OptiBond Universal（OPB; Kerr, Brea, CA, USA）,

Introduction

Prime&Bond Universal（PBU; Dentsply Sirona, Char-The three-step bonding system consists of etching,（UBQ; Kuraray Noritake Dental, Tokyo, Japan）. The

priming, and bonding procedures. Due to the demand

characteristics of each adhesive are as follows. Regard-

for simpler, more user-friendly, and less technique-sen-ing the functional monomers, OPB contains GPDM,

sitive adhesives, manufacturers have developed new

PBU contains MDP and PENTA, and UBQ contains

adhesives used in one-step procedures1,2）. Recently, one-

MDP. Additionally, PBU and UBQ contain a hydrophiliclotte, NC, USA）, and Clearfil Universal Bond Quick ER

step self-etch adhesive（SEA）systems called“multi-

amide monomer, and only UBQ contains NaF. Table 1

mode adhesive”or“universal adhesive”were released.

provides a detailed description of the respective adhe-With these adhesives, clinicians can choose one of threesives and application protocols. In this study, caries-freestrategies（etch and rinse, selective enamel etch, or self-

human third molars were collected according to theetch modes）, depending on the clinical application, ren-ethical protocol No. 2013-022 approved by the Human3）

dering the clinical procedure more user-friendly .

Research Ethics Committee of Tokyo Medical and Den-

These bonding agents are also indicated to be used astal University. The teeth were stored in 0.1％ thymolsilane for glass ceramics and primers for metal alloys

solution at 4℃, and used within six months afterand polycrystalline ceramics. This is why these adhe-extraction.

sives are called“universal adhesives” . Universal adhe-2 ．Microtensile bond strength（μTBS）testingsives are a complex mixture of hydrophilic and hydro-

1 ）Tooth specimen preparation

4）

phobic compounds, water, and solvents such as ethanol

The outline of the μTBS test is schematically pre-and acetone, which can lead to poor adhesion and dete-sented in Fig. 1. Thirty human third molars wererioration of the bonding interface5）.

selected for this part of the study. For each tooth, theTo evaluate the bonding interface, not only the bondcoronal portion was removed to expose a flat mid-coro-strength test but also morphological evaluation is often

nal dentin surface using a low-speed water-cooled dia-

conducted. Primarily, a scanning electron microscopemond saw（Isomet; Buehler, Lake Bluff, IL, USA）. The（SEM）examination provides valuable information for

exposed dentin surface was ground for 30 s with #600-

adequately understanding the mechanism of adhesive

grit silicon carbide（SiC）paper under running water toadhesion to teeth. SEM studies after acid and base chal-produce standardized smear layers.

lenges revealed the formation of an“acid-base resistant2 ）Bonding and restorative procedureszone（ABRZ）”adjacent to the hybrid layer when dentin

Thirty teeth were randomly divided into three

was treated with a SEA system6）. However, there havegroups（10 teeth per group）by type of adhesive. Eachbeen few comparative studies7） on universal adhesives

tooth was bonded according to the manufacturer’s

7）

after acid and base challenges. In one study , a univer-

instructions（Table 1）. After application, the specimens

sal adhesive was employed to evaluate the bonding

were air-dried for 5 s and cured with a light-emittinginterface.

diode（LED）light-curing unit（VALO LED Curing Light,

Therefore, the present study elucidated the den-high-power mode at 1,400 mW/cm2, Ultradent, Southtin-bonding performance of three kinds of universal

Jordan, UT, USA）for 10 s. Then, the resin composite

adhesive by the bond strength test and morphological（Clearfil AP-X, shade A2, Kuraray Noritake Dental）

observation under SEM. The null hypothesis of thiswas built to a thickness of 2 mm and light-cured forstudy was that the material components of universal

40 s using the light-curing unit. This operation was per-adhesives do not influence the durability of dentin

formed twice to build a resin composite block with abonding.

total thickness of 4 mm. After storage in 37℃ distilledwater for 24 h, all bonded teeth were sectioned into

Materials and Methods

beam-shaped specimens（surface area: 1.0×1.0 mm）
using Isomet. Nine specimens were obtained from the

1 ．Materials used in this study
This study used three kinds of universal adhesive,

center of each bonded tooth（9 beams per tooth）.
The beam-shaped specimens in each group were ran-ODEP Vol. 1, No. 1

90
Surface preparation#600 SiC

Bonding（OPB，
UBQ，PBU）
composite

24 hour storage
in distilled water at 37°C

Human molar
Thermal cyclingtransfer time 5 s

Sectioning

5°C 30 s
Fig. 155°C 30 s

Prime & Bond
Universal

Clearfil Universal
Bond Quick ER

Clearfil Majesty
ES Flow

Crosshead speed
1 mm/minFailure mode
analysis

Materials used in this study
Hydrophilicamidemonomers

HEMA

NaF

Application time

OPB was applied for
20 s with agitation,

Code

Composition

OPB

GPDM, HEMA, dimethacrylate,acetone, ethanol, glycerol,

―

✓

―

PBU

Phosphoric acid modified acrylate
resin（MDP, PENTA）, multifunctional acrylate, bifunctional acrylate（amide monomer）, acidic acrylate,isopropanol, water, initiator

✓

―

―

UBQ

MDP, HEMA, Bis-GMA, hydrophilic
amide monomers, colloidal silica,
silane coupling agent, sodium fluoride, dl-camphorquinone, ethanol,
water

Resin composite
Clearfil AP-X

SEM

Schematic of specimen preparation methodology for μTBS
Table 1

Universal adhesive
OptiBond
Universal

μTBS

then air-dried for 5 s.
PBU was applied for
20 s with agitation,then air-dried for 5 s.

✓✓✓
UBQ was applied and（＜0.1％） immediately air-dried
with no time lapse
between steps.

Bis-GMA, TEGDMA, silanated glass filler, silanated silica filler, silanated colloidal silica, di-camphorquinone, catalysts, accelerators, pigments
Silanate barium glass filler, silanate colloidal filler, TEGDMA, hydrophobic aromatic dimethacrylate, di-camphorquinone, accelerators, pigments

GPDM：glycero-phosphate dimethacrylate, HEMA：2-hydroxyethyl methacrylate, MDP：10-methacryloyloxydecyl dihydrogen phosphate, PENTA：dipentaerythritol penta acrylate monophosphate, TEGDMA：triethylene glycol dimethacrylate, BisGMA：2,2 bis［4-（2-hydroxy-3-methacrylyloxy-propoxy）-phenyl］propane.

domly divided into three subgroups, and exposed to 0,

device（K178-08, Tokyo Giken, Tokyo, Japan）. After

5,000 or 10,000 thermal cycles（n＝30, for each sub-thermal cycling, each specimen was individually bonded

group）. The beams were cycled between two water

to a tensile testing jig using cyanoacrylate adhesivebaths at 5℃ and 55℃ with a dwell time of 30 s in each

（Model Repair II Blue, Dentsply-Sankin, Tokyo, Japan）

bath and a transfer time of 5 s using a thermal cycling

mounted in a tabletop testing machine（EZ-SX, Shi-Dec, 2021

Effects of Universal Adhesives on Dentin Adhesive and Acid-base Resistance91

Bonding（OPB，UBQ，
PBU）
composite#600 SiC

Dentin disc
Human molar

Sectioning
2 mm

24 hour storage
in tap water
Embeddingat 37°C

Acid－base challenge

PolishingFig. 2

Schematic of specimen preparation methodology for SEM

madzu, Kyoto, Japan）
and subjected to the μTBS test at

flowable resin composite（Clearfil Majesty ES Flow,a crosshead speed of 1.0 mm/min（Fig. 1）. The μTBS

shade A2, Kuraray Noritake Dental）was placed

was derived by dividing the imposed force at the time

between pairs of prepared dentin disks and light-curedof fracture by the bonded area（mm2）. Statistical differ-

to make a dentin disk sandwich. The bonded specimens

ences were examined by multiple comparisons using

were stored in distilled water at 37℃ for 24 h. Subse-

t-tests with Bonferroni correction（α＝0.05）in IBM

quently, the specimens were sectioned vertically at the

SPSS Statistics for Windows, version 22.0（IBM,dentin-adhesive interface and embedded in epoxy resin

Armonk, NY, USA）.

（Epoxicure 2 Resin, Buehler）. Each specimen was first

After the μTBS test, the fractured specimens werestored in 100 ml of a buffered demineralizing solution,observed under SEM（JSM-IT100, JEOL, Tokyo, Japan）

containing 2.2-mmol/l CaCl2, 2.2-mmol/l NaH2PO4, andto characterize their failure modes. Fractured speci-

50-mmol/l acetic acid adjusted to pH 4.5, for 90 min tomens were classified as:（1）C-D: cohesive failure increate an artificial secondary caries challenge. The

dentin with＞80％ of total bonding area,（2）I-DA: inter-

specimens were then immersed in 5％ NaOCl（pH＝facial failure between dentin and adhesive with＞80％10.5‒12.9）, which was prepared from 10％ sodium hypo-of total bonding area,（3）C-A: cohesive failure in adhe-chlorite solution（Yoneyama Yakuhin Kogyo, Osaka,

sive with＞80％ of total bonding area,（4）Mix: mixed

Japan）, using an ultrasonic cleaner for 20 min in an

failure. Statistical differences were examined by Fisher’s

attempt to remove any demineralized dentin collagenexact test and Chi-square test（α＝0.05）in IBM SPSS

fibrils, and rinsed with running water for 30 s. The

Statistics for Windows, version 22.0.

edge of the dentin-adhesive interface could be easily

3 ．SEM obser vation of the adhesive-dentin interface after acid-base challenge

damaged during specimen polishing. To prevent this, aself-cure adhesive resin, Super-Bond C&B（Sun Medical,

The sample preparation for SEM examination of theMoriyama, Japan）, was applied to the surface without

ABRZ is illustrated in Fig. 2. Thirty human third

any surface preparation8）. The specimens were sec-

molars were randomly divided into three groups（10tioned perpendicular to the dentin-adhesive interface,teeth per group）. Each tooth was sectioned to obtainand reduced to approximately 2 mm thickness, thentwo 2 mm-thick dentin disks using Isomet. The dentin

polished with diamond pastes（Struers A/S, Copenha-

surfaces were ground with #600-grit SiC paper undergen, Denmark）down to 0.25μm particle size. The pol-running water to standardize the smear layers, and inished surfaces were etched with an argon-ion beam

the same way as the μTBS test, one of the three adhe-（EIS-200ER, Elionix, Tokyo, Japan）at 1 kV, 0.2 mA/cm2

sives was applied. Following the adhesive application, afor 30 s to bring the hybrid layer into sharp relief. The

ODEP Vol. 1, No. 1

92
Table 2Group

Microtensile bond strength（Mean±SD, MPa）
Thermal cycle
0

5,000

10,000OPB

67.5±13.2A

PBU

62.4±8.2Aa

58.6±10.0B
57.5±10.6Bab

47.4±11.6C
52.8±9.8CDb

UBQ

66.3±9.4A

58.7±6.4Bc

54.3±13.1Dc

N＝30, Mean and standard deviation（SD）of microtensile
bond strength values（MPa）.
Values within the same thermal cycle stage, marked withthe same large superscripts, are not significantly different（p＞0.05）.
Values within the same material, marked with the same
small superscripts, are not significantly different（p＞0.05）.

specimens were then gold-sputter coated, and morphological changes to the dentin-adhesive interface due tothe acid-base challenge were observed under SEM9,10）.

Results
1 ．Microtensile bond strength（μTBS）
The μTBS values and failure modes are shown in
Tables 2 and 3. Compared to each adhesive, the μTBS

Table 3Group

Failure mode analysis

Thermal cycle

OPB

I-DA

C-A

Mix

a

19a

1

4

a

5,000

11

a

0

a

8

a

11a

10,000

9a

3a

6a

12a

a

0

a

a

14a

1

a

a

12

4a

0

a

a

8a

0
5,000
10,000

UBQ

C-D

a

0

PBU

Failure mode

6

10
13

a

15

a

6
7

0
5,000

a

11
16a

a

2
2a

a

0
3a

17a
9a

10,000

15a

1a

6a

8a

C-D：cohesive failure in dentin with ＞80％ of total
bonding area, I-DA：interfacial failure between dentin
and adhesive with ＞80％ of total bonding area, C-A：
cohesive failure in adhesive with ＞80％ of total bonding area, Mix：mixed failure.
Numbers marked with the same small superscripts are
not significantly different（p＞0.05）.

ence in all groups.
2 ．SEM obser vations of ultra-structural features
after acid-base challenge

test results showed that the bond strength of the UBQ

Typical interface morphologies after the acid-basegroups was significantly higher than that of the OPB

challenge are shown in Fig. 3. An outer lesion（OL）,

groups after the 10,000 thermal cycle challenge（p＜created by the dissolution of dentin due to the acid-base

0.001）. Regarding the bond strength of the 0 and 5,000

challenge, was observed in the three groups. The depth

cycles groups, there were no significant differencesof OL ranged from 15 to 20μm in all specimens. ABRZ

among the three adhesive groups. In the OPB groups,

was observed in all groups. The zone was approxi-

the bond strength of the 0 cycle group was significantly

mately less than 1μm thick. Moreover, a funnel-shaped

higher than those of the 5,000 and 10,000 cycles groups

erosion was observed at the junction of the dentin and（p＜0.001）. Moreover, the bond strength of the 5,000

bonding layer in all groups. A funnel-shaped erosion

cycles group was significantly higher than that of thewas clearly observed in OPB and PBU. On the other

10,000 cycles group（p＜0.001）. In the UBQ groups, thehand, a funnel-shaped erosion was observed in UBQ, butbond strength of the 0 cycle group was significantly

not clearly.

higher than those of the 5,000 and 10,000 cycles groups（p＜0.001）. However, there was no statistically signifi-

Discussion

cant difference in the bond strength between the 5,000
and 10,000 cycles groups（p＞0.05）. In the PBU groups,

For 5,000 thermal cycle challenges, μTBS results

there was no statistically significant difference in theshowed different trends among the three groups. Of thebond strength between the 0 and 5,000 cycles groups

three universal adhesives, only the OPB group showed（p＞0.05）. Moreover, there was no difference in thea statistically significant deterioration in μTBS at bothbond strength of the 5,000 and 10,000 cycles groups

5,000 and 10,000 cycles. Both UBQ and PBU contain（p＞0.05）. Meanwhile, the bond strength of the 0 cycle

multifunctional hydrophilic amide monomers. A multi-group was significantly higher than that of the 10,000

functional hydrophilic amide monomer is more hydro-

cycles group（p＜0.001）. Regarding the failure mode

philic before polymerization and more hydrophobic

observation, there was no statistically significant differ-

after polymerization than previously used hydrophilicDec, 2021

Effects of Universal Adhesives on Dentin Adhesive and Acid-base Resistance93

Fig. 3 Representative SEM images of failure site（a）C-D：cohesive failure in the dentin with ＞80％ of total bonding area, magnification of 80×.（b）I-DA：interfacial
failure between dentin and adhesive with ＞80％ of total bonding area, magnification of 80×.（c）C-A：cohesive failure inadhesive with ＞80％ of total bonding area, magnification of 80×.（d）Mix：mixed failure, magnification of 80×.（e）C-D,
magnification of 500×.（f）I-DA, magnification of 500×.（g）C-A, magnification of 500×.

Fig. 4 SEM images of dentin-adhesive interface after acid-base challenge, magnification of 3,500×
OL：outer lesion, R：resin composite, A：adhesive layer, D：dentin, ABRZ：acid-base resistant zone, OPB：SEM imageof OptiBond Universal, PBU：SEM image of Prime&Bond Universal, UBQ：SEM image of Clearfil Universal Bond QuickER.
ABRZ was observed in all groups. In OPB and PBU a funnel-shaped erosion was clearly observed at the junction of thedentin and bonding layer, whereas a funnel-shaped erosion was not clearly observed in UBQ.

monomers, and also has excellent mechanical proper-

exhibits hydrophilic properties, leading to water uptake

ties11）. Therefore, these characteristics of hydrophilic

after polymerization13）. Thus, HEMA-containing SEAs

amide monomer might contribute to the bond strength

are more hydrophilic14）, exhibit higher water sorption15）

after the thermal cycles of UBQ and PBU.

and show an accelerated reduction in the mechanicalFurthermore, the PBU group results showed no sta-

properties of the adhesive layer during water stor-

tistically significant difference between 0 cycle andage16）. Additionally, some studies12,17‒20） reported that

5,000 cycles（p＞0.05）. In this study, only PBU does not

HEMA reduces the long-term durability of adhesive.

contain 2-hydroxyethyl methacrylate（HEMA）. The

From the results of earlier studies and this study, ithydrophilic monomer HEMA has been commonly incor-

could be inferred that PBU showed more resistance toporated into adhesive formulations to improve dentin

the thermal cycle test than other adhesives.

bond strength because it enhances wetting of the den12）

tin subsurface . However, HEMA after polymerization

For dentin bonding, the hybridization theory has
been widely accepted, in which a resin monomer pene-ODEP Vol. 1, No. 1

94

trates into demineralized dentin and polymerizes invent and water is clinically difficult35）. Therefore, thesitu, resulting in the formation of a hybrid layer, which

degree of polymerization of universal adhesives is gen-is an intermingling of resin with the collagen fibers oferally inferior to that of 2SEAs5）. These properties of

21,22）

the dentin

23）

first reported

universal adhesives could make a vulnerable region

a new zone adjacent to the visible hybrid layer in 2SEA

adjacent to the bonding interface, resulting in the for-characterized by the resistance to acid and base chal-mation of a funnel-shaped erosion after the acid-baselenges, which was identified from a part of the hybrid

challenge. In contrast, in this experiment, a fun-layer by argon etching. This zone indicated higher

nel-shaped erosion was clearly observed in OPB andmicrohardness than the hybrid layer, and was charac-

PBU, but not in UBQ. Previous studies of ABRZ36,37）

terized by tooth-like structures under SEM observa-

reported the formation of a slope shape, sloping upwardtion. In 2004, Tsuchiya et al.

23）. Hence, it is different from the conventional

to the end of the outer lesion, without a funnel-shaped

hybrid layer and caries-inhibition zone in fluoride-re-

erosion observed when using SEAs containing NaF.

leasing materials. It was therefore named“ABRZ,”and

The formation of a slope shape indicates improved acidis considered to play an important role in preventing

resistance. From these reports, NaF might improve thesecondary caries, sealing restoration margins, and pro-acid-base resistance adjacent to the bonding inter-21）

moting restoration durability . However, ABRZ forma-

face36,37）. Meanwhile, the formation of a funnel-shapedtion has been confirmed only in SEA systems, but not in

erosion indicates that the acid-base resistance of theetch and rinse systems

21,24）. Furthermore, it was

region directly below the ABRZ is reduced. In thisreported that ABRZ is not formed depending on theexperiment, a funnel-shaped erosion was not clearly25）

type of functional monomer . In this experiment, allobserved in UBQ, probably because the decrease ingroups formed ABRZ. Previous studies 7,26） demon-

acid-base resistance was compensated for by the inclu-

strated that ABRZ was formed using 2SEA containing

sion of NaF. From these results, the null hypothesis of

MDP or GPDM. In this study, it was newly confirmed

this study was rejected.

that GPDM contained in universal adhesives also forms
ABRZ. Meanwhile, some researches7,27）reported that it

Conclusion

was difficult to detect the hybrid layer at the interfaceof 1SEA, even though the hybrid layer was detected

After thermal cycling, the bond strength testwith 2SEA. Similarly, the hybrid layer was not clearly

appeared to be influenced by the hydrophilic amide

observed using universal adhesives in this study. Somemonomer（PBU and UBQ）or HEMA（OPB and UBQ）.

studies7,26,28） reported that even if ABRZ is formed, a

For SEM observation after the acid-base challenge,

funnel-shaped erosion adjacent to ABRZ indicates a vul-

ABRZ formation was detected using universal adhe-nerable region to acid-base attacks. It was reported

sives containing MDP（PBU and UBQ）or GPDM（OPB）.

that a universal adhesive that contains MDP creates

Moreover, a funnel-shaped erosion adjacent to ABRZ

ABRZ with a funnel-shaped erosion7,26）, and that a fun-was not clearly observed in a universal adhesive con-nel-shaped erosion was not observed in 2SEA contain-

taining NaF（UBQ）, but was clearly observed in others. In contrast to 2SEA, in this

（OPB and PBU）
. Therefore, the results of this study

study, all groups formed a funnel-shaped erosion,suggest that the durability of the bonding between uni-regardless of the type of functional monomer.

versal adhesives and dentin depends on material com-ing MDP or GPDM

7,26）

Universal adhesives contain hydrophobic monomers,

ponents.

hydrophilic monomers, water, and solvents in one bottle. Hence, universal adhesives are a complex mixture

Conflict of Interestof hydrophilic and hydrophobic compounds and water,This study was approved by the Conflict of Interest Committee of Tokyo Medical and Dental University（C-A20201201）.

so they contain solvents such as ethanol or acetone tomaintain miscibility29,30）. Meanwhile, solvent and water
evaporation is essential because residual solvent andwater adversely affect the polymerization of the adhesive12,31‒34）. However, complete evaporation of the sol-Dec, 2021

Effects of Universal Adhesives on Dentin Adhesive and Acid-base Resistance95

chemical composition of contemporary dental adhesives.

References
1）Seitoku E, Hoshika S, Ikeda T, Abe S, Tanaka T, Sano H.
Bonding performance of a hydrophilic amide monomer
containing adhesive to occlusal and cervical dentin.
Materials（Basel）2020; 13: 4727.
2）Van Meerbeek B, De Munck J, Yoshida Y, Inoue S, Vargas M, Vijay P, Van Landuyt K, Lambrechts P, Vanherle G. Buonocore memorial lecture. Adhesion toenamel and dentin: Current status and future challenges. Oper Dent 2003; 28: 215‒235.
3）Perdigão J, Kose C, Mena-Serrano AP, De Paula EA, Tay
LY, Reis A, Loguercio AD. A new universal simplified
adhesive: 18-month clinical evaluation. Oper Dent 2014;
39: 113‒127.
4）Giannini M, Makishi P, Ayres AP, Vermelho PM, Fronza
BM, Nikaido T, Tagami J. Self-etch adhesive systems: A
literature review. Braz Dent J 2015; 26: 3‒10.
5）Sato T, Takagaki T, Hatayama T, Nikaido T, Tagami J.
Update on enamel bonding strategies. Front Dent Med
2021; 2: 666379.
6）Nikaido T, Takagaki T, Sato T, Burrow MF, Tagami J.
Fluoride-releasing self-etch adhesives create thick
ABRZ at the interface. Biomed Res Int 2021; 2021:
9731280.
7）Guan R, Takagaki T, Matsui N, Sato T, Burrow MF,
Palamara J, Nikaido T, Tagami J. Dentin bonding performance using Weibull statistics and evaluation of acidbase resistant zone formation of recently introduced
adhesives. Dent Mater J 2016; 35: 684‒693.
8）Inoue G, Nikaido T, Foxton RM, Tagami J. The acid-baseresistant zone in three dentin bonding systems. Dent
Mater J 2009; 28: 717‒721.
9）Vicheva M, Sato T, Takagaki T, Baba Y, Ikeda M, Burrow MF, Nikaido T, Tagami J. Effect of repair systems
on dentin bonding performance. Dent Mater J 2021; 40:903‒910.
10）Nakamoto A, Sato T, Matsui N, Ikeda M, Nikaido T,
Burrow MF, Tagami J. Effect of fluoride mouthrinse
and fluoride concentration on bonding of a one-step selfetch adhesive to bovine root dentin. J Oral Sci 2019; 61:
125‒132.
11）Kuno Y, Hosaka K, Nakajima M, Ikeda M, Klein Junior
CA, Foxton RM, Tagami J. Incorporation of a hydrophilic amide monomer into a one-step self-etch adhesive
to increase dentin bond strength: Effect of application
time. Dent Mater J 2019; 38: 892‒899.
12）Van Landuyt KL, Snauwaert J, De Munck J, Peumans M,
Yoshida Y, Poitevin A, Coutinho E, Suzuki K, Lambrechts P, Van Meerbeek B. Systematic review of the

Biomaterials 2007; 28: 3757‒3785.
13）Ito S, Hashimoto M, Wadgaonkar B, Svizero N, Carvalho
RM, Yiu C, Rueggeberg FA, Foulger S, Saito T, Nishitani Y, Yoshiyama M, Tay FR, Pashley DH. Effects ofresin hydrophilicity on water sorption and changes inmodulus of elasticity. Biomaterials 2005; 26: 6449‒6459.
14）Yiu CK, King NM, Carrilho MR, Sauro S, Rueggeberg
FA, Prati C, Carvalho RM, Pashley DH, Tay FR. Effectof resin hydrophilicity and temperature on water sorption of dental adhesive resins. Biomaterials 2006; 27:1695‒1703.
15）Malacarne J, Carvalho RM, de Goes MF, Svizero N,
Pashley DH, Tay FR, Yiu CK, Carrilho MR, de Oliveira
Carrilho MR. Water sorption/solubility of dental adhesive resins. Dent Mater 2006; 22: 973‒980.
16）Takahashi M, Nakajima M, Hosaka K, Ikeda M, Foxton
RM, Tagami J. Long-term evaluation of water sorption
and ultimate tensile strength of hema-containing/-free
one-step self-etch adhesives. J Dent 2011; 39: 506‒512.
17）Van Landuyt KL, Snauwaert J, De Munck J, Coutinho E,
Poitevin A, Yoshida Y, Suzuki K, Lambrechts P, Van
Meerbeek B. Origin of interfacial droplets with one-stepadhesives. J Dent Res 2007; 86: 739‒744.
18）Itoh S, Nakajima M, Hosaka K, Okuma M, Takahashi M,
Shinoda Y, Seki N, Ikeda M, Kishikawa R, Foxton RM,
Tagami, J. Dentin bond durability and water sorption/solubility of one-step self-etch adhesives. Dent Mater J
2010; 29: 623‒630.
19）Shinoda Y, Nakajima M, Hosaka K, Otsuki M, Foxton
RM, Tagami J. Effect of smear layer characteristics on
dentin bonding durability of HEMA-free and HEMAcontaining one-step self-etch adhesives. Dent Mater J
2011; 30: 501‒510.
20）Van Landuyt KL, Snauwaert J, Peumans M, De Munck
J, Lambrechts P, Van Meerbeek B. The role of HEMA
in one-step self-etch adhesives. Dent Mater 2008; 24:1412‒1419.
21）Matsui N, Takagaki T, Sadr A, Ikeda M, Ichinose S,
Nikaido T, Tagami J. The role of MDP in a bonding
resin of a two-step self-etching adhesive system. Dent
Mater J 2015; 34: 227‒233.
22）Gale MS, Darvell BW. Thermal cycling procedures forlaboratory testing of dental restorations. J Dent 1999;
27: 89‒99.
23）Tsuchiya S, Nikaido T, Sonoda H, Foxton RM, Tagami J.
Ultrastructure of the dentin-adhesive interface afteracid-base challenge. J Adhes Dent 2004; 6: 183‒190.
24）Inoue G, Tsuchiya S, Nikaido T, Foxton RM, Tagami J.
Morphological and mechanical characterization of the
acid-base resistant zone at the adhesive-dentin interface
of intact and caries-affected dentin. Oper Dent 2006; 31:

ODEP Vol. 1, No. 1

96
466‒472.

sives and chemically-cured or dual-cured composites.

25）Li N, Nikaido T, Takagaki T, Sadr A, Makishi P, Chen J,
Tagami J. The role of functional monomers in bonding

Part iii. Effect of acidic resin monomers. J Adhes Dent

to enamel: Acid-base resistant zone and bonding performance. J Dent 2010; 38: 722‒730.
26）Sato T, Takagaki T, Matsui N, Hamba H, Sadr A,
Nikaido T, Tagami J. Morphological evaluation of theadhesive/enamel interfaces of two-step self-etchingadhesives and multimode one-bottle self-etching adhesives. J Adhes Dent 2016; 18: 223‒229.
27）Inoue G, Nikaido T, Sadr A, Tagami J. Morphologicalcategorization of acid-base resistant zones with
self-etching primer adhesive systems. Dent Mater J
2012; 31: 232‒238.
28）Nikaido T, Takagaki T, Sato T, Burrow MF, Tagami J.
The concept of super enamel formation―Relationshipbetween chemical interaction and enamel acid-baseresistant zone at the self-etch adhesive/enamel interface. Dent Mater J 2020; 39: 534‒538.
29）De Munck J, Van Landuyt K, Peumans M, Poitevin A,
Lambrechts P, Braem M, Van Meerbeek B. A criticalreview of the durability of adhesion to tooth tissue:
Methods and results. J Dent Res 2005; 84: 118‒132.
30）Van Landuyt KL, Peumans M, De Munck J, Lambrechts
P, Van Meerbeek B. Extension of a one-step self-etchadhesive into a multi-step adhesive. Dent Mater 2006;
22: 533‒544.
31）Suh BI, Feng L, Pashley DH, Tay FR. Factors contributing to the incompatibility between simplified-step adhe-32）Frankenberger R, Strobel WO, Lohbauer U, Krämer N,
Petschelt A. The effect of six years of water storage on
resin composite bonding to human dentin. J Biomed
Mater Res B Appl Biomater 2004; 69: 25‒32.
33）Tay FR, Pashley DH, Suh BI, Hiraishi N, Yiu CK. Water
treeing in simplified dentin adhesives―déjà vu? Oper
Dent 2005; 30: 561‒579.
34）Loguercio AD, Loeblein F, Cherobin T, Ogliari F, Piva E,
Reis A. Effect of solvent removal on adhesive propertiesof simplified etch-and-rinse systems and on bondstrengths to dry and wet dentin. J Adhes Dent 2009; 11:213‒219.
35）Ikeda T, De Munck J, Shirai K, Hikita K, Inoue S, Sano
H, Lambrechts P, Van Meerbeek B. Effect of evaporation of primer components on ultimate tensile strengthsof primer-adhesive mixture. Dent Mater 2005; 21: 1051‒
1058.
36）Kirihara M, Inoue G, Nikaido T, Ikeda M, Sadr A, Tagami J. Effect of fluoride concentration in adhesives onmorphology of acid-base resistant zones. Dent Mater J
2013; 32: 578‒584.
37）Kakiuchi Y, Takagaki T, Ikeda M, Sato T, Matsui N,
Nikaido T, Burrow MF, Tagami J. Evaluation of MDP
and NaF in two-step self-etch adhesives on enamel
microshear bond strength and morphology of the adhesive-enamel interface. J Adhes Dent 2018; 20: 527‒534.

2003; 5: 267‒282.

Dec, 2021

97

Case Report

ODEP 1（1）：97～104，2021

A Case of Non-surgical Root Canal Treatment Using an Operating
Microscope on Type Ⅲ-b Dens Invaginatus
Masashi YAMADA1,5, Kento ODAKA2,5, Norio KASAHARA3,5,
Satoru MATSUNAGA4,5, Yoshiki TAMIYA1, Ryo SAKO1
and Masahiro FURUSAWA1
1

Department of Endodontics, Tokyo Dental College

2

Department of Oral and Maxillofacial Radiology, Tokyo Dental College

3

Histology and Developmental Biology and Clinical Pathophysiology, Tokyo Dental College
4

Department of Anatomy, Tokyo Dental College

5

Oral Health Science Center, Tokyo Dental College

Abstract
Dens invaginatus is a developmental malformation, in which there is an infolding of enamel into dentin. It
is prevalent in maxillary lateral incisors, with a relatively high incidence of 0.3‒10％. However, the morphology is complex and therefore its diagnosis and treatment are difficult. We report a case of endodontic treatment of dens invaginatus with a root apex lesion by non-surgical root canal treatments using an operating
microscope to preserve the pulp.
Case report: The patient was a 23-year-old male. He had no spontaneous pain but had an abscess of themaxillary left lateral incisor. The pulp was diagnosed as reversible pulpitis. Transient pain occurred uponcold stimulation. Percussion and palpation pain were noted. The periodontal pocket depth was ≤3 mm allaround the tooth. Dental X-rays showed invagination in the central part of the root canal and there were
pathways of communication with the periodontal tissue at the root apex. CBCT was taken to understand theroot canal morphology. Oehlers type Ⅲ-b dens invaginatus was diagnosed, with apical periodontitis due toinfection via the invagination despite a normal tooth pulp morphology. Therefore, root canal treatment of theinvagination was performed using a microscope and pulp preservation was anticipated. Both oral and written
informed consent were obtained from the patient prior to treatment, and a full explanation of the research
purpose was given. A manual stainless-steel file was then used to explore the route through the pseudo-root
canal and was passed through it, and the root canal length was measured with an electronic apex locator.
After confirming that the root canal length was correct, the working length was set at a point 0.5 mm
shorter than the root canal length. Preparation was performed using Ni-Ti rotary files. After final irrigationof the root canal, filling was performed with MTA cement. The entrance of the invagination was sealed withan adhesive resin.
Course: The clinical symptoms disappeared after one year. The case was diagnosed as“healing”due to thediminishing apical radiolucency on dental X-ray and CBCT.
Discussion: In this case, we were able to magnify the invagination to the minimum necessary by using an
operating microscope. The use of MTA cement enabled tight sealing and protection of the pulp, which may
have resulted in good healing.
Conclusions: An operating microscope may be useful in the treatment of dens invaginatus.
Key words: dens invaginatus, Oehlers’ type Ⅲ-b, CBCT, operating microscope
Corresponding author: Masashi YAMADA, Department of Endodontics, Tokyo Dental College, 2‒19‒18, Kanda-Misakicho,
Tokyo 101‒0061, Japan
TEL: ＋81‒3‒6380‒9136, E-mail: myamada21@tdc.ac.jp
Received for Publication: September 6, 2021/Accepted for Publication: October 11, 2021
DOI: 10.11471/odep.2021‒009

ODEP Vol. 1, No. 1

98week. On inspection, the maxillary left lateral incisor

Introduction

had a wide tooth crown and a lingual-side composite
resin filling, and the gingiva in the apical region was

Dens invaginatus, also known as tooth within a tooth,

reddened（Figure 1-a）. In terms of clinical symptoms,

is a condition in which there is invagination of thecold stimulation elicited transient pain, and the tooth

enamel and dentin of a tooth crown into the pulp cav-

exhibited a normal pulp reaction in comparison with1）

ity ; its mean prevalence is reported to be 0.3‒10％.

that of the neighboring teeth, but percussion pain, apicalAround 90％ of reported cases have involved the maxil-

palpation, and some mobility were evident. The peri-

2）

lary lateral incisors , and it has been shown to be moreodontal pocket depth was ≤3 mm all around the tooth

frequent in particular ethnic groups3‒5）.

and some degree of tooth mobility was evident. On a

Although numerous different classification methods

panoramic X-ray image that the patient brought fromhave been reported, Oehler’s classification, which isthe previous doctor, the maxillary left lateral incisor

based on the depth of incursion of the invagination andshowed root hypertrophy, and its appearance suggested

its positional relationship with the cementoenamel junc-

dens invaginatus. Dental X-ray imaging was therefore

tion
（CEJ）
, is generally used5）. Studies of the prevalence

performed for further investigation（Figure 1-b）, and anof different classes have not identified any racial differ-invagination that reached the apex of the root wasences, with typeⅠ being the most frequent and type Ⅲ

found in the center of the tooth; there was no obvious

6,7）. This is also the case in Japa-

caries-like radiolucency in the crown, but an apicalnese patients8）. Type Ⅲ has two subtypes, with invagi-

radiolucency was apparent around the hypertrophic

nations that communicate laterally with the periapicalroot apex. On the basis of these findings, Oehler’s typetissue designated as type Ⅲ-a and those that communi-

Ⅲ-b dens invaginatus was diagnosed, with apical peri-cate via the apical foramen as type Ⅲ-b. Because theseodontitis due to infection via the invagination despite ainvaginations are completely separate from the pulp

normal tooth pulp morphology. When embarking on

cavity, they may cause large apical lesions even in vitaltreatment, additional cone-beam computed tomography

teeth. Such cases are difficult to treat and diagnose, but（CBCT）was performed to evaluate the shape of thecases in which the tooth pulp has been conserved byinvagination and its positional relationship with thesuccessful treatment of the invagination in the samelesion（Figure 2）. On the CBCT images, the enamel wasway as an infected root canal（pseudo-root canal）haveinvaginated from the incisal tip toward the apex andthe most uncommon

9）been reported . This report describes an attempt tocommunicated with the periodontal tissue, so that theconserve the tooth pulp of a type Ⅲ-b dens invaginatus

invagination appeared similar to a root canal（pseu-

with an apical lesion that nevertheless exhibited a vital

do-root canal）. In part of the interior of this pseudo-root

reaction by performing non-surgical root canal treat-canal, the enamel was partially cut off and the dentin

ment using an operating microscope.

was exposed. The apical radiolucency included theopening of the invagination, and a part of the buc-Case Reportcal-side cortical bone defect was evident in the sagittal
section. The slice data acquired by CBCT were used toPatient: 23-year-old man

create a three-dimensional（3D）reconstruction image

Chief complaint: Unable to bite due to swelling of theand a 3D model for use in treatment planning. The 3D

gingiva of the left upper front teeth

model was printed with a 3D printer（Objet260 Connex,

Location: Maxillary left lateral incisor

Stratasys, Eden Prairie, MN, USA）using transparent

General medical history: Nothing of note

biocompatible resin（MED610, Stratasys）（Figure 3）.

History of present illness: The patient was examined

On the 3D reconstruction image, there was no obvi-

for root canal treatment of a malformed tooth at theous communication between the main root canal andrequest of his previous dentist. A hole had appeared on

the pseudo-root canal. On the 3D-printed model, thethe lingual side of the maxillary left lateral incisor two

pulp cavity was observed to surround the pseudo-root

months before and this had been filled. The patient said

canal in a C-shape. The pulpal horn was present up to athat the gingiva had been swollen since the previous

comparatively high position. Based on these findings, a

Dec, 2021

RCT of Dens Invaginatus Using a Microscope

a

99

b

Fig. 1 Pre-treatment images of maxillary left lateral incisor（a）Intraoral photographs showing a wide crown of the lateral incisor.（b）Dental X-ray image showing an
enlarged root and an invagination penetrating to the root apex.

c

d

a

b
e

Fig. 2 Pre-treatment CBCT images of maxillary left lateral incisor
The cortical bone of the buccal side is missing.
（a）Frontal section.（b）Sagittal section.（c）（d）Transverse,
sections.

treatment plan was formulated consisting of non-surgi-

understood them, consented to take part. Treatment

cal treatment under surgical microscopic observation,

was then started in compliance with the ethical regula-with the aim of preserving the pulp by treating thetions of Tokyo Dental College（Tokyo Dental College

pseudo-root canal as a root canal.

Ethics Committee approval no. 999）.

In line with the Declaration of Helsinki, the purposeof the study and the treatment plan were explainedorally and in writing to the patient, who, having fully

Treatment: Root canal treatment of the pseudo-root
canal.
The composite resin closing the invagination was

ODEP Vol. 1, No. 1

100

a

b

c

d

Fig. 3 3D construction images of maxillary left lateral incisors（a）3D construction image.（b）3D printer output buccal-side model.（c）3D printer output distal-sidemodel.（d）3D printer output mesial-side model.

a

b

c

Fig. 4 Enlargement of the dens invaginatus opening and negotiation（a）After enlargement of the dens invaginatus opening.（b）Drainage from the pseudo-root canal.
（c）Dental X-ray with a trial file at the root canal length.

removed with a diamond turbine bur, after which thefirm that this length was in fact correct. The working

enamel of the invagination was drilled away. A manual

length was then designated as a point 0.5 mm proximal

stainless-steel file was then used to explore the routeto the root canal length（Figure 4）. The hardness of thethrough the pseudo-root canal and was passed throughenamel in the upper part of the pseudo-root canalit. There was a purulent discharge from the pseu-

meant that the entrance had not been sufficiently

do-apex, which was aspirated using a root canal vac-enlarged, and this was therefore further enlarged using

uum. When no more pus was discharged, an electronica diamond-coated ultrasonic tip. Because exploration

apex locator was used to measure the length of the rootwith the manual file showed that the inner surface of

canal. A file was inserted into the root canal up to thisthe pseudo-root canal felt softer to drill, it was enlarged

measured length, and a dental X-ray was taken to con-to #60 with a Ni-Ti rotary file（Hi-Flex CM, ColteneDec, 2021

RCT of Dens Invaginatus Using a Microscope

a

b

d

101

c

e

f

g

Fig. 5 Intraoperative images of maxillary left lateral incisor（a）Before enlargement of the dens invaginatus opening.（b）After enlargement of the dens invaginatus
opening with ultrasonic instruments.（c）Enlargement of the pseudo-root canal.（d）After cleaning the pseudo-root canal.（e）（f）,
Filling the pseudo-root canal with MTA cement.（g）After filling the pseudo-root canal.

Whaledent, Cuyahoga Falls, OH, USA）
. However,tal X-rays showed that the radiolucency had decreased

because there were some hard parts of the inner sur-in size. One year after filling, there was no change inface of the pseudo-root canal, it was thought that it

symptoms, but on dental X-rays, a radiolucency was

might not have been sufficiently enlarged, and chemical

still present on the inner curve（Figure 6）. CBCT scan-cleaning was therefore performed more thoroughlyning was performed to evaluate whether the buc-than during normal root canal treatment. Passive ultra-

cal-side bone had healed. On CBCT images, a radiolu-

sonic irrigation（PUI）was performed using 2.5％ NaOCl

cency was still evident on the inner curve of the invagi-

and an ultrasonic tip. A final cleaning of the inside ofnation, but since regeneration of the buccal-side cortical

the pseudo-root canal was conducted using 2.5％ NaOClbone was observed, it was considered to be healing

and 17％ ethylenediaminetetraacetic acid（EDTA）,（Figure 7）.

after which it was dried with a root canal vacuum. The
pseudo-root canal was filled with mineral trioxideaggregate（MTA）cement. A rubber stopper was fitted

Discussion

to a root canal filling plugger, and incremental filling

In the treatment of type Ⅲ dens invaginatus with an

was performed while taking care that nothing escaped

apical lesion, if the tooth pulp exhibits a vital reaction, itfrom the pseudo-apex into the periapical tissue. The use

is reportedly possible to conserve the pulp by treatingof MTA as a root filling material was approved by thethe pseudo-root canal10‒12）. However, issues of concern

Ethics Committee of Tokyo Dental College Suidobashi

in the treatment of pseudo-root canals include:（1）theHospital（approval no. SH-41）. After pseudo-root canal

inner surface of the invagination is covered withfilling, the entrance to the invagination was closed withenamel and irregular in shape;（2）in parts of the innercomposite resin, and dental X-rays were then taken

surface, dentin is exposed, and the pulp is closely adja-（Figure 5）.

cent; and（3）the pseudo-apical foramen is wide open10）.
In the present case, the following two issues were also

Clinical Course

considered for the three risk factors. The first issue
was the appropriate instrument to use for cleaning out

Three months after non-surgical root canal treat-

the pseudo-root canal. The instruments reported inment, the clinical symptoms had disappeared, and den-studies can be broadly divided into three types: manualODEP Vol. 1, No. 1

102

a

b

Fig. 6 Post-treatment X-rays of the maxillary left lateral incisors（a）Three months after root canal filling.（b）Twelve months afterroot canal filling.

a

e

b

f

c

d

g

h

Fig. 7 Pre-treatment and post-treatment images（a）Pre-treatment intraoral photographs.（b）‒（d）Pre-treatment CBCT images.（e）Post-treatment intraoral photographs. Loss of sinus tract was observed.（f）（‒ h）
Post-treatment CBCT images. Bone regeneration
was observed at the root apex and buccal side.

drilling instruments, rotary drilling instruments, andwidely reported9,11）. In the present case, the enamel at

ultrasound drilling instruments. Manual drilling instru-

the invagination entrance was hard, and the inner sur-ments enable the torque during enlargement to be con-face of the pseudo-root canal was irregular. Hard areas

trolled by the operator, preventing instrument fracture

were therefore encountered at various points on theand enabling safe treatment, and their use has been

inner surface, and a manual instrument could not beDec, 2021

RCT of Dens Invaginatus Using a Microscope

103

used because the drilling sensation was difficult to dis-due to the fact that we were able to select the appro-

tinguish. With respect to rotary drilling instruments,

priate filling method based on the diagnosis using thethere have been some reports on the use of Gates-Glid-

3D model in advance20）. Another report has also noted

den drills in combination with manual drilling instru-

that, after root canal treatment of a type Ⅲ dens invag-

9,13）

ments

, but if the enamel of the inner surface of theinatus with similar morphology, a radiolucency on theinvagination is hard, then drilling with them is also diffi-

inner curve of the root persisted for a long time despite

cult. The use of a Ni-Ti rotary file has also been

the absence of clinical symptoms21）. In that report by

14）. In the present patient,

Sübay et al., the radiolucency on the inner curve of the

because most of the inner surface of the pseudo-root

root persisted for 3 years after pseudo-root canal treat-canal was covered with enamel, the areas of advanced

ment with no evident clinical symptoms. They there-

caries had softened and could be drilled away, but thefore performed surgical curettage, which showed

other parts could not be sufficiently drilled. Rotary drill-

regeneration of the bone. In the present case as well, aing instruments may be captured by the enamel insidepersistent bone defect at the same site was still present

the pseudo-root canal, and if torsion stress is generated,

after 18 months. However, it is possible that healing

they may break10）. Their use thus requires caution andmay have been delayed because of the large size of thecare. Ultrasound instruments are reportedly capable ofbone defect, or that it may never have been filled withefficiently enlarging and removing the invagination15）,bone at all because of the persistence of soft tissue

and they can enable sufficient debridement when usedfrom the time of eruption. We will continue to monitor

in conjunction with cleaning16）; in the present case, one

this case carefully, including the possibility of future

was useful for drilling out the invagination entrance andsurgical intervention.

reported in a few cases

debridement during the PUI of the inside of the pseudo-root canal. Ultrasound instruments can also be used

Conclusion

under surgical microscopic observation, which enablessafe, minimally invasive treatment. We therefore con-This case demonstrated the usefulness of an operat-

sider ultrasound instruments to be ideal for use ining microscope for the non-surgical, pulp-conserving

cleaning dens invaginatus. The second issue was which

treatment of type Ⅲ-b dens invaginatus.

material and method to use for filling the pseudo-root
canal. In this case, the pseudo-root canal was closely
adjacent to the pulp based on the diagnosis using a 3D

The authors state that there are no conficts of interest
related to this study.

model, and pulp irritation and infection during enlargement and filling were both concerns. It is well known
that MTA cement causes little damage to pulp, and it isrecommended as a pulp capping agent17）, reportedly
providing good long-term closure and therapeutic outcomes18）. We therefore considered that the use of MTA
cement was appropriate to prevent pulp irritation andinfection. Because the pseudo-apex was wide open, we
had to consider the possibility that the filling agent
might escape into the periapical tissue. In the conventional root canal filling method using gutta-percha, lateral and vertical condensation filling techniques havebeen reported 11）, but in both techniques, the filling
agent may escape, and caution is therefore required.
Good results have been reported from the use of MTA
cement even when the apical foramen is wide open18,19）,and it may therefore be suitable for filling pseudo-root
canals. In addition, the success of the treatment was

References
1）Hallett GEM. Incidence, nature and clinical significanceof palatal invagination in the maxillary incisor teeth.
Proc R Soc Med 1953; 46: 491‒499.
2）Hamasha AA, Alomari QD. Prevalence of dens invaginatus in Jordanian adults. Int Endod J 2004; 37: 307‒310.
3）Cho SY, Ki Y, Chu V, Chan J. Concomitant developmental dental anomalies in Chinese children with dens evaginatus. Int J Paediatr Dent 2006; 16: 247‒251.
4）Shi S, Duan X, Shao J, Duan Q, Peng S. Dens invaginatus
in ancient Chinese teeth of 2,000 years ago. Anat Rec（Hoboken）2013; 296: 1628‒1633.
5）Oehlers FA. Dens invaginatus. I. Variations of the invagination process and associated anterior crown forms.
Oral Surg Oral Med Oral Pathol 1957; 10: 1204‒1218.
6）Ridell K, Mejàre I, Matsson L. Dens invaginatus: a retrospective study of prophylactic invagination treatment.

104
Int J Paediatr Dent 2001; 11: 92‒97.

ODEP Vol. 1, No. 1
Andrabi SM. Double dens invaginatus with multiple7）Capar ID, Ertas H, Arslan H, Tarim Ertas E. A retrospective comparative study of cone-beam computed

canals in a maxillary central incisor: Retreatment andtomography versus rendered panoramic images in identifying the presence, types, and characteristics of dens
invaginatus in a Turkish population. J Endod 2015; 41:473‒478.
8）Gotoh T, Kawahara K, Imai K, Kishi K, Fujiki Y. Clinicaland radiographic study of dens invaginatus. Oral Surg
Oral Med Oral Pathol 1979; 48: 88‒91.
9）Schwartz SA, Schindler WG. Management of a maxillarycanine with dens invaginatus and a vital pulp. J Endod
1996; 22: 493‒496.
10）Bishop K, Alani A. Dens invaginatus. Part 2: clinical,

15）De Smit A, Demaut L. Nonsurgical endodontic treatment of invaginated teeth. J Endod 1982; 8: 506‒511.
16）Girsch WJ, McClammy TV. Microscopic removal of dens
invaginatus. J Endod 2002; 28: 336‒339.
17）Mente J, Geletneky B, Ohle M, Koch MJ, Friedrich Ding
PG, Wolff D, Dreyhaupt J, Martin N, Staehle HJ, Pfefferle T. Mineral trioxide aggregate or calcium hydroxide direct pulp capping: an analysis of the clinical treatment outcome. J Endod 2010; 36: 806‒813.
18）Pace R, Giuliani V, Nieri M, Di Nasso L, Pagavino G.
Mineral trioxide aggregate as apical plug in teeth withnecrotic pulp and immature apices: a 10-year caseseries. J Endod 2014; 40: 1250‒1254.
19）Torabinejad M, Chivian N. Clinical applications of mineral trioxide aggregate. J Endod 1999; 25: 197‒205.
20）Kfir A, Telishevsky-Strauss Y, Leitner A, Metzger Z.
The diagnosis and conservative treatment of a complex
type 3 dens invaginatus using cone beam computed

radiographic features and management options. Int
Endod J 2008; 41: 1137‒1154.
11）Fristad I, Molven O. Root resorption and apical breakdown during orthodontic treatment of a maxillary lateral incisor with dens invaginatus. Endod Dent Traumatol 1998; 14: 241‒244.
12）Tsurumachi T. Endodontic treatment of an invaginatedmaxillary lateral incisor with a periradicular lesion anda healthy pulp. Int Endod J 2004; 37: 717‒723.
13）Pitt Ford HE. Peri-radicular inflammation related todens invaginatus treated without damaging the dental
pulp: a case report. Int J Paediatr Dent 1998; 8: 283‒286.
14）Zoya A, Ali S, Alam S, Tewari RK, Mishra SK, Kumar A,

managing complications. J Endod 2015; 41: 1927‒1932.

tomography（CBCT）and 3D plastic models. Int Endod
J 2013; 46: 275‒288.
21）Sübay RK, Kayataş M. Dens invaginatus in an immature
maxillary lateral incisor: a case report of complex endodontic treatment. Oral Surg Oral Med Oral Pathol Oral
Radiol Endod 2006; 102: e37‒41.

Dec, 2021

105

Submission Guidelines for ODEP
1．ODEP aims to develop conservative dentistry（operative dentistry, endodontology, and periodontology）
throughthe publication of research and reviews on the following topics:（1）General dental medicine, clinical practice,and education on conservative dentistry; and（2）Conservative dentistry.
2．Papers are categorized into the following four types:（1）Original articles（reports on unique research discoveries）;（2）Reviews（discussions on research questions and objectives to indicate future directions, or summariesof the contents of existing papers to propose new ideas）;（3）Mini reviews（concise summaries of recent topics;
mini reviews include papers awarded with prizes）; and（4）Case and clinical reports（analysis of clinicalrecords useful for dental care practice and development of the field of conservative dentistry）. Reviews andmini reviews are categorized into the following:（1）Papers requested by the editorial board; and（2）Submitted
papers.
3．Original articles and case and clinical reports are limited to the following:（1）Papers that have not been publishedin other journals;（2）Papers that are not presently submitted to another journal; and（3）Papers that are notpresently scheduled for publication.
4．Acceptance or rejection of papers is determined through peer review（except for papers requested by the editorial board）.
5．Submitted papers should be concisely written in English.
6．In principle, original articles should be organized as follows:（1）Abstract;（2）Introduction;（3）Materials andMethods;（4）Results;（5）Discussion;（6）Conclusion;（7）References;（8）Figure legend;（9）Figures and Tables.
In principle, papers other than original articles should conform to the style format of original articles.
7．In principle, ODEP is published once a year in December. In addition, special issues are published when appropriate.
8．The Japanese Society of Conservative Dentistry provides a certain amount of support for the listing fees of
papers. For cases in which the lead author is not a member of the Japanese Society of Conservative Dentistry,
such support is not provided. For cases in which the lead author is a member, but the co-authors include a nonmember, partial support is provided. The cost to be borne by the authors is determined based on the numberof non-members. The costs for figures, tables and photographs, for dispatching and offprints, and for creating
the J‒STAGE registration data are borne by the authors. In the case of papers requested by the editorial
board, such costs are exempted.
9．Date of submission is the date when the submitted manuscript arrives at the secretariat of the Japanese Societyof Conservative Dentistry. Date of acceptance is the date when the reviewers determine that the submitted
manuscript can be published.
10．The listing order is the order of acceptance. A certification of publication will be issued upon request.
11．Manuscripts are to be submitted via the Japanese Society of Conservative Dentistry’s website, e-mail, or postal
mail. Manuscripts submitted for publication should be addressed to the secretariat of the Japanese Society ofConservative Dentistry.
12．In principle, authors can proofread their manuscript a maximum of two times. Extensive changes, additions, ordeletions made to the contents of the manuscript cannot be accepted. Proofs should be returned by the designated date. If the authors do not need to proofread their manuscript, they should mention this on the left sideof the cover page.
13．The copyrights for articles published in ODEP belong to the Japanese Society of Conservative Dentistry.
14．Matters not mentioned in these guidelines will be independently determined by the editorial board.
Submission of your manuscript for publication must conform to the following“Submission Guidance”as well as“Submission Guidelines.”ODEP Vol. 1, No. 1

106

The publishing charge is 10,000 yen for a Journal page including tables and figures. Extra charges for such as
figures and tables preparation, color printing of photographs will also be paid by the authors. If authors do not
pay publishing charge, the article may be retracted.

Submission Guidance（applied as of the Issue 1 of Vol. 2）
Manuscript organization
1．In principle, original articles should be organized into the following sections:（1）cover page;（2）abstract;（3）Main text（Introduction, Materials and Methods, Results, Discussion, and Conclusion）;（4）References; and（5）Figure and table captions. Page numbering should start with the cover page. In principle, manuscripts, such as
reviews or case reports, other than original articles should be organized in the same format as that of originalarticles.
2．Manuscript sections
1） Title: The title should concisely describe the contents of the manuscript. The subtitle should also clearly
describe the contents, and should not consist of only numbers.
2） Introduction: The introduction should clearly describe the background, novelty, purpose, and significance ofthe study.
3） Materials and Methods: This section should provide detailed information on the materials, equipment, or
methods used along with clear instructions so that the experiments can be reproduced by others. Parameter settings, number of specimens, extraction methods, statistics processing, and others should comply withthe purpose of the study.
4） Results: This section should simply present the findings without bias or interpretation. Measurement results
should show characteristic values including mean values and standard deviations.
5） Discussion: This section should carefully consider the materials and methods, results, and others referring
to relevant literature. Please note that it should not be overly assertive and should avoid off-topic points.
The discussion should stay focused on the study purpose and not digress into a general discussion.
6） Conclusion: The section should precisely summarize the results obtained and relate them to the purpose of
the study and hypothesis as presented in the Introduction.
3．Manuscripts should be prepared using A4-size paper. The suggested length of each typed page is 80 alphanumeric characters per line×25 lines per page using a 12-point font. Top/bottom/left/right margins should beapproximately 25 mm.

Non-Japanese names and places should be in their original names..

4．For the manuscript style, refer to the latest issue of the journal.
Ethics Code
1．Each report on the result of clinical research（clinical trial or observational research）or research involving any
specimen collected from a human body must include a clear statement that it was approved by the head of theaffiliated institution or by the research ethics review board assigned by the institution’s head, in order toexpressly indicate that the research was conducted in compliance with all applicable guidelines and laws,
including the Declaration of Helsinki and the medical research guidelines, etc. issued by the Ministry of Health,Labour and Welfare including the following:
1）Ethical Guidelines for Life Science and Medical Research Involving Human Subjects;
2）Guidelines for Gene Therapy and Other Clinical Research; and3）Clinical Trials Act.
2．Each report on the result of research on, or a case concerning, regenerative medicine technology, etc. as definedDec, 2021

107

in the Act on the Safety of Regenerative Medicine must include a clear statement that the technology was provided to the patients in compliance with the aforementioned Act.
3．Each report on a case of a therapeutic method involving any off-label drug or device, or any pharmaceutical,
medical device, regenerative medicine, or other related product not domestically approved must include a clear
statement that the use was approved by the committee concerned（research ethics review board, review
board for unapproved new drugs, etc.）at the affiliated institution.
4．In each instance of publication of an academic paper, all personal information must be thoroughly protected sothat none of the research subjects（patients）can be identified from it.
5．In each instance where a patient’s clinical photo or X-ray image is included in an academic paper for publication,a clear statement must be provided that the consent of each such patient（or a parent, guardian, or proxy if thepatient is a minor or in case it is otherwise difficult to obtain consent from the patient）was duly obtained.
6．Each report on the result of research involving animal subjects must include a clear statement that the research
was approved by the animal experiment committee, etc. at the affiliated institution.
Cover page
1．The title, authors’ names, institutional affiliations, and corresponding author’s contact address should be centered,with a new line for each item, on the cover page.
2．The title should be in upper and lowercase letters, where the first letter of each word is uppercased and theremaining letters are lowercased. Articles, prepositions, conjunctions, and commonly used technical terms arelowercased. For hyphenated compound words, the letters following the hyphen should be lowercased.
3．The corresponding author’s contact details should include the following information: one author’s name, institutional affiliation, postal address, telephone and fax numbers, and e-mail address.
Abstract
1．The abstract should be a maximum of 400 words organized into four sections with the following headings: Purpose, Methods, Results, and Conclusion. Approximately three keywords should be placed at the end of theabstract.
2．Contributors should put considerable effort into preparing the abstract as it may determine whether or not thereader continues with the manuscript. When necessary, abstracts should be checked by a native English
reviewer（preferably with expertise in dental medicine）.
Main text
1．Introduction, Materials and Methods, Results, Discussion, and Conclusion are the main headings and are notnumbered.
2．Subparagraphs should be numbered in the following order: 1. 2. 3. …; 1）2）3）…;（1）（2）（3）…; ① ② ③ …;
and a. b. c. …
3．English characters should be written in the following manner:
1） Generally, only the last name should be used to indicate a person.
2） When the name of a product or manufacturer must be in the original language, the first letter of each word
should be uppercased and the remaining letters lowercased.
In principle,“generic name（product name, company name, city［state in the case of U.S.］, country）”
shouldbe used in English manuscripts. Trademark and registration symbols ® and TM are not required.
3） Regarding common nouns in German or Latin, the first letter should be uppercased and the remaining letters lowercased. For common nouns in English and French, all letters should be lowercased.
4） Regarding binominal nomenclature, the first letter of the genus should be uppercased and the remaining
letters lowercased. The names of all genera and species should be italicized. When the same genus
appears frequently, it is acceptable to replace the name with the initial after the first use.

ODEP Vol. 1, No. 1

108

Example: Streptococcus mutans → S. mutans
5） For nouns that must be in their original language, other than German, Latin, English or French, all lettersshould be lowercased, except for commonly used technical terms.
4．In principle, SI units are used for measurements.
5．Any conflicts of interests（COI）must be declared after the conclusions. When there is no COI, the statement“The authors declare no conflict of interest related to this paper”should be included.
6．Acknowledgments for all sources supporting this study including grant funds should be added after of COI.
References
1．References must be listed at the end of the main text, and numbered in the same order as they appear in thetext.
2．In the main text, a cited reference should appear with a superscript numeral and closing parenthesis. When two
references are cited, a comma should be used to separate them; more than two references should be connectedby an en dash between the first and last numeral.
Examples:“by authors3）”,“…is reported7,8）”,“previous studies10‒15）show”
3．Examples of Reference
a ．Journal articles
Number）Last name and first name of all authors with a comma separating each author. Title of paper. Name ofjournal and publishing year in the Christian era; Volume number: Inclusive page numbers of paper.
Example:1）Clark AB, Erickson D, Hamilton FG. Tensile bond strength and modulus of elasticity of several composite resins. J Dent
Res 1992; 37: 618‒621.

b ．Book
Number）Author（co-authors）. Title of book. First/last volume. Edition. Publisher’s name: Publisher’s location（City）; Publishing year in the Christian era. Cited pages.
Example:2）Phillips RW. Skinner’s science of dental materials. 9th ed. WB Saunders: Philadelphia; 1991. 219‒221.

c ．Book with co-authors
Number）Contributor’s name. Title of contributed article. Name of editor（editor-in-chief）. Book title. First/lastvolume. Edition. Publisher’s name: Publisher’s location（City）; Publishing year in the Christian era.
Cited pages.
Example:3）Torneck CD. Dentin-pulp complex. Ten Cate AR. Oral histology. 5th ed. Mosby: St. Louis; 1998. 150‒196.

For cases in which each author’s contribution is not separately indicated, the author’s name and the title of thecontributed article should not be listed.
Number）Name of editor（editor-in-chief）. Title of book. First/last volume. Edition. Publisher’s name: Publisher’slocation（City）; Publishing year in the Christian era. Cited pages.
d ．Other writing styles
・Abstracts of scientific meeting
Number）Presenter（all presenters should be cited. A comma should be used to separate the authors in the caseof co-presentations.）. Title of abstract. Name of journal and publishing year in the Christian era; Volume number: Inclusive page numbers, abstract number.
Example:4）Marais JT. Cleaning efficacy of a new root canal irrigation material. J Dent Res 1998; 77: 669, Abst. No. 300.

・ Journal articles in press
In principle, the same style as that of a regular journal article should be used; however, when the inclusive page
numbers are not known, they can be omitted. The statement“in press”should be shown at the end.

Dec, 2021

109

Example:5）Sato K. Effect of toothbrushes on gingival abrasion. J Periodont Res 1994; 29: in press.

・Electronic journal
In principle, the same style as that of a regular journal article should be used; however, when the inclusive page
numbers are not known, the DOI and other indicators should be shown. When papers are published in electronic journals before printing, the statement［Epub ahead of print］should be shown after the publishing year
and month.
Example:6）Sunada N, Ishii R, Shiratsuchi K, Shimizu Y, Tsubota K, Kurokawa H, Miyazaki M. Ultrasonic measurement of theeffects of adhesive application and power density on the polymerization behavior of core build-up resins. Acta Odontol Scand; doi: 10.3109/00016357.2011.654252

・Internet website
Page publisher. Title of page. URL address.（Access date）
Example:7）World Health Organization. Continuous improvement of oral health in the 21st century. http://www.who.int/oral_
health/en/（cited 2005. 10. 1）

4．In principle, journal names should be abbreviated in accordance with the format used by the journal.
Figures and Tables
1．Figures, photographs, and tables are categorized into figures and tables, and then numbered. Paper size shouldbe A4, and each figure and table should be printed on a separate page. The numbers allocated for the figures
and tables should be consistent with those referred to in the text.
2．Figures and tables should be accompanied by explanations that are easily understandable. Explanations of figures/tables are presented as captions and explanations for tables/tables should be presented as footnotes.
3．If authors wish to have the figures color-printed, color data should be attached; if authors wish to have the figures
printed in black and white, black-and-white data should be attached.
Notes on creating imaging data:
・ The jpg data format should be used if possible.
・ Image size should correspond to the layout; image resolution should be at least 300 dpi for photographs andat least 1,200 dpi for line drawings.
4．Figures printed in black and white can be uploaded as color pictures on J‒STAGE; authors should request thisat the time of submission. The cost to create color data for J‒STAGE uploading is to be borne by the authors.
In a case where color figures are uploaded only on J‒STAGE, new captions cannot be added since it would
make the J‒STAGE content different from the original content published in the journal. When selecting color/
black-and-white figures, the authors themselves should carefully check that the original figure captions are sufficient.
Sending manuscript intended for publication
1．The manuscript（cover page, abstract, main text, references, and figure and table captions are created as onefile）should be formatted as a Microsoft Office Word（hereafter“Word”）document.
2．Figures should be provided in jpg or pdf format.
3．Tables should be provided in Microsoft Office Excel, jpg, pdf, or Word format.
4．File titles should be as follows:“author name”_“university name（name after department is not needed）”_“manuscript/figure/table/submission form”_“.filename extension（indicating file type）”.
Example）Nihon Tarou_Nihon University/manuscript.docx; Nihon Taro_Nihon University_Figure.jpg; Nihon
Taro_Nihon Univesity_Table.xlsx; Nihon Taro_Nihon University_submission form.pdf
Also, pdf files with embedded fonts for all contents are acceptable for submission. In such cases, file names

ODEP Vol. 1, No. 1

110should be as follows: Nihon Taro_Nihon University_comprehensive manuscript.pdf
5．E-mail title（subject）should be“Submitted papers for ODEP”.
6．Submitted papers should be sent to the e-mail address of Oral Health Association of Japan: hensyu6@kokuhoken.
or.jp. For safety, also send as a CC to hensyu5@kokuhoken.or.jp.
7．When e-mail submission is difficult for such reasons as the file size is too large, submission via an FTP server
and other methods are acceptable. In such cases, the intent of submission should be informed by e-mail. In thee-mail, information regarding the site and other methods for downloading the file should be provided.
8．The submission form of ODEP provided on the Japanese Society of Conservative Dentistry’s home page（http://www.kokuhoken.or.jp/exterior/ jscd/fileform/）can be used for submission.

Reprographic Reproduction
The Japanese Society of Conservative Dentistry authorized Japan Academic Association For Copyright
Clearance（JAC）to license our reproduction rights of copyrighted works. If you wish to obtain permissionsof these rights, please refer to the homepage of JAC（http://www.jaacc.org/en/）and confirm appropriateorganizations to request permission.

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